U.S. patent application number 15/905685 was filed with the patent office on 2018-07-05 for human antibodies to pd-1.
The applicant listed for this patent is Regeneron Pharmaceuticals, Inc.. Invention is credited to Elena BUROVA, Ella IOFFE, Andrew J. MURPHY, Nicholas J. PAPADOPOULOS, Gavin THURSTON.
Application Number | 20180185668 15/905685 |
Document ID | / |
Family ID | 52462456 |
Filed Date | 2018-07-05 |
United States Patent
Application |
20180185668 |
Kind Code |
A1 |
PAPADOPOULOS; Nicholas J. ;
et al. |
July 5, 2018 |
HUMAN ANTIBODIES TO PD-1
Abstract
The present invention provides antibodies that bind to the
T-cell co-inhibitor programmed death-1 (PD-1) protein, and methods
of use. In various embodiments of the invention, the antibodies are
fully human antibodies that bind to PD-1. In certain embodiments,
the present invention provides multi-specific antigen-binding
molecules comprising a first binding specificity that binds to PD-1
and a second binding specificity that binds to an autoimmune tissue
antigen, another T-cell co-inhibitor, an Fc receptor, or a T-cell
receptor. In some embodiments, the antibodies of the invention are
useful for inhibiting or neutralizing PD-1 activity, thus providing
a means of treating a disease or disorder such as cancer or a
chronic viral infection. In other embodiments, the antibodies are
useful for enhancing or stimulating PD-1 activity, thus providing a
means of treating, for example, an autoimmune disease or
disorder.
Inventors: |
PAPADOPOULOS; Nicholas J.;
(LaGrangeville, NY) ; MURPHY; Andrew J.;
(Croton-on-Hudson, NY) ; THURSTON; Gavin;
(Briarcliff Manor, NY) ; IOFFE; Ella; (Bronx,
NY) ; BUROVA; Elena; (Mount Kisco, NY) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Regeneron Pharmaceuticals, Inc. |
Tarrytown |
NY |
US |
|
|
Family ID: |
52462456 |
Appl. No.: |
15/905685 |
Filed: |
February 26, 2018 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
14603776 |
Jan 23, 2015 |
|
|
|
15905685 |
|
|
|
|
61930576 |
Jan 23, 2014 |
|
|
|
62014181 |
Jun 19, 2014 |
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 3/10 20180101; C07K
2317/76 20130101; A61K 2039/545 20130101; A61P 31/14 20180101; A61P
35/00 20180101; A61P 37/04 20180101; C07K 16/2818 20130101; A61K
45/06 20130101; A61P 31/20 20180101; A61P 1/14 20180101; A61P 17/14
20180101; A61P 31/12 20180101; A61P 21/04 20180101; A61P 31/18
20180101; C07K 2317/92 20130101; A61P 1/18 20180101; A61P 7/04
20180101; A61P 7/06 20180101; C07K 2317/21 20130101; A61P 17/06
20180101; A61P 37/06 20180101; A61P 1/16 20180101; A61P 29/00
20180101; A61K 39/3955 20130101; A61P 19/02 20180101; A61P 37/08
20180101; A61P 17/00 20180101; A61N 5/10 20130101; C07K 16/2827
20130101; A61P 25/00 20180101; A61P 17/04 20180101; A61K 2039/505
20130101; A61P 37/02 20180101; A61P 1/04 20180101; C07K 2317/31
20130101; A61K 39/3955 20130101; A61K 2300/00 20130101 |
International
Class: |
A61N 5/10 20060101
A61N005/10; C07K 16/28 20060101 C07K016/28; A61K 45/06 20060101
A61K045/06; A61K 39/395 20060101 A61K039/395 |
Claims
1-56. (canceled)
57. An isolated nucleic acid molecule comprising a nucleic acid
sequence encoding a heavy chain variable domain region (HCVR) of an
antibody that binds programmed death 1 (PD-1), wherein the HCVR
comprises a heavy chain CDR1 (HCDR1) comprising SEQ ID NO: 164, a
heavy chain CDR2 (HCDR2) comprising SEQ ID NO: 166, and a heavy
chain CDR3 (HCDR3) comprising SEQ ID NO: 168.
58. The nucleic acid molecule of claim 57, wherein the HCVR
comprises the amino acid sequence of SEQ ID NO: 162.
59. The nucleic acid molecule of claim 57, wherein the HCVR
comprises an HCDR1 encoded by the nucleotide sequence of SEQ ID
NO:163, an HCDR2 encoded by the nucleotide sequence of SEQ ID NO:
165, and an HCDR3 encoded by the nucleotide sequence of SEQ ID NO:
167.
60. The nucleic acid molecule of claim 57, wherein the nucleic acid
molecule comprises the nucleotide sequence of SEQ ID NO: 161 or a
substantially identical sequence having at least 95% homology
thereof.
61. The nucleic acid molecule of claim 57, wherein the nucleic acid
molecule comprises the nucleotide sequence of SEQ ID NO: 161.
62. An isolated nucleic acid molecule comprising a nucleic acid
sequence encoding a light chain variable region (LCVR) of an
antibody that binds PD-1, wherein the LCVR comprises a light chain
CDR1 (LCDR1) comprising SEQ ID NO: 172, a light chain CDR2 (LCDR2)
comprising, SEQ ID NO: 174, and a light chain CDR3 (LCDR3)
comprising SEQ ID NO: 176.
63. The nucleic acid molecule of claim 62, wherein the LCVR
comprises SEQ ID NO: 170.
64. The nucleic acid molecule of claim 62, wherein the LCVR
comprises an LCDR1 encoded by the nucleotide sequence of SEQ ID NO:
171, an LCDR2 encoded by the nucleotide sequence of SEQ ID NO: 173,
and an LCDR3 encoded by the nucleotide sequence of SEQ ID
NO:175.
65. The nucleic acid molecule of claim 62, wherein the nucleic acid
molecule comprises the nucleotide sequence of SEQ ID NO: 169 or a
substantially identical sequence having at least 95% homology
thereof.
66. The nucleic acid molecule of claim 62, wherein the nucleic acid
molecule comprises the nucleotide sequence of SEQ ID NO: 169.
67. An expression vector comprising: (a) a nucleic acid molecule
comprising a nucleic acid sequence encoding a HCVR of an antibody
that binds PD-1, wherein the HCVR comprises a HCDR1 comprising SEQ
ID NO: 164, a HCDR2 comprising SEQ ID NO: 166, and a HCDR3
comprising SEQ ID NO: 168; and/or (b) a nucleic acid molecule
comprising a nucleic acid sequence encoding a LCVR of an antibody
that binds PD-1, wherein the LCVR comprises a LCDR1 comprising SEQ
ID NO: 172, a LCDR2 comprising SEQ ID NO: 174, and a LCDR3
comprising SEQ ID NO: 176.
68. An isolated host cell comprising the expression vector of claim
67.
69. The host cell of claim 68, wherein the host cell is a mammalian
cell or a prokaryotic cell.
70. The host cell of claim 68, wherein the host cell is a Chinese
Hamster Ovary (CHO) cell or an Escherichia coli (E. coli) cell.
71. A method of producing an anti-PD-1 antibody or antigen-binding
fragment thereof, the method comprising growing the host cell of
claim 68 under conditions permitting production of the antibody or
antigen-binding fragment thereof, wherein said host cell comprises
both a nucleic acid molecule comprising a nucleic acid sequence
encoding said HCVR and a nucleic acid molecule comprising a nucleic
acid sequence encoding said LCVR.
72. The method of claim 71, further comprising formulating the
antibody or antigen-binding fragment thereof as a pharmaceutical
composition comprising an acceptable carrier.
73. A composition comprising a first nucleic acid molecule and a
second nucleic acid molecule; wherein the first nucleic acid
molecule comprises a nucleic acid sequence encoding a HCVR of an
anti-PD-1 antibody that comprises a HCDR1 comprising SEQ ID NO:
164, a HCDR2 comprising SEQ ID NO: 166, and a HCDR3 comprising SEQ
ID NO: 168; and wherein the second nucleic acid molecule comprises
a nucleic acid sequence encoding a LCVR of an anti-PD-1 antibody
that comprises a LCDR1 comprising SEQ ID NO: 172, a LCDR2
comprising SEQ ID NO: 174, and a LCDR3 comprising SEQ ID NO:
176.
74. The composition of claim 73, wherein the HCVR comprises SEQ ID
NO: 162, and wherein the LCVR comprises SEQ ID NO: 170.
75. An isolated nucleic acid molecule encoding an antibody or
antigen-binding fragment thereof, which specifically binds PD-1,
wherein the antibody or antigen-binding fragment comprises a heavy
chain variable region (HCVR) comprising three heavy chain CDRs
(HCDR1, HCDR2 and HCDR3) from SEQ ID NO: 162, and a light chain
variable region (LCVR) comprising three light chain CDRs (LCDR1,
LCDR2 and LCDR3) from SEQ ID NO: 170.
76. The nucleic acid molecule of claim 75, wherein: (a) the HCDR1
comprises SEQ ID NO: 164; (b) the HCDR2 comprises SEQ ID NO: 166;
(c) the HCDR3 comprises SEQ ID NO: 168; (d) the LCDR1 comprises SEQ
ID NO: 172; (e) the LCDR2 comprises SEQ ID NO: 174; and (f) the
LCDR3 comprises SEQ D NO: 176.
77. The nucleic acid molecule of claim 76, wherein the HCVR
comprises SEQ ID NO: 162.
78. The nucleic acid molecule of claim 76, wherein the LCVR
comprises SEQ ID NO: 170.
79. The nucleic acid molecule of claim 76, wherein the HCVR
comprises SEQ ID NO: 162 and the LCVR comprises SEQ ID NO: 170.
80. An expression vector comprising the nucleic acid molecule of
claim 75.
81. An isolated host cell comprising the expression vector of claim
80.
82. The host cell of claim 81, wherein the host cell is a mammalian
cell or a prokaryotic cell.
83. The host cell of claim 81 wherein the host cell is a Chinese
Hamster Ovary (CHO) cell or an Escherichia coli (E. coli) cell.
84. A method of producing an anti-PD-1 antibody or antigen-binding
fragment thereof, comprising growing the host cell of claim 81
under conditions permitting production of the antibody or fragment,
and recovering the antibody or fragment so produced.
85. The method of claim 84, further comprising formulating the
antibody or antigen-binding fragment thereof as a pharmaceutical
composition comprising an acceptable carrier.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This is a Continuation of U.S. patent application Ser. No.
14/603,776, filed Jan. 23, 2015, which claims the benefit under 35
U.S.C. .sctn. 119(e) of U.S. provisional application Nos.
61/930,576, filed on Jan. 23, 2014; and 62/014,181, filed on Jun.
19, 2014, the disclosures of each herein incorporated by reference
in their entireties.
FIELD OF THE INVENTION
[0002] The present invention is related to human antibodies and
antigen-binding fragments of human antibodies that specifically
bind to the immunomodulatory receptor programmed death-1 (PD-1),
and therapeutic and diagnostic methods of using those
antibodies.
STATEMENT OF RELATED ART
[0003] Programmed death-1 (PD-1) (also called CD279) is a 288 amino
acid protein receptor expressed on activated T-cells and B-cells,
natural killer cells and monocytes. PD-1 is a member of the
CD28/CTLA-4 (cytotoxic T lymphocyte antigen)/ICOS (inducible
co-stimulator) family of T-cell co-inhibitory receptors (Chen et al
2013, Nat. Rev. Immunol. 13: 227-242). The primary function of PD-1
is to attenuate the immune response (Riley 2009, Immunol. Rev. 229:
114-125). PD-1 has two ligands, PD-ligand1 (PD-L1) and PD-L2. PD-L1
(CD274, B7H1) is expressed widely on both lymphoid and non-lymphoid
tissues such as CD4 and CD8 T-cells, macrophage lineage cells,
peripheral tissues as well as on tumor cells, virally-infected
cells and autoimmune tissue cells. PD-L2 (CD273, B7-DC) has a more
restricted expression than PD-L1, being expressed on activated
dendritic cells and macrophages (Dong et al 1999, Nature Med.).
PD-L1 is expressed in most human cancers, including melanoma,
glioma, non-small cell lung cancer, squamous cell carcinoma of head
and neck, leukemia, pancreatic cancer, renal cell carcinoma, and
hepatocellular carcinoma, and may be inducible in nearly all cancer
types (Zou and Chen 2008, Nat. Rev. Immunol. 8: 467-77). PD-1
binding to its ligands results in decreased T-cell proliferation
and cytokine secretion, compromising humoral and cellular immune
responses in diseases such as cancer, viral infection and
autoimmune disease. Blockade of PD-1 binding to reverse
immunosuppression has been studied in autoimmune, viral and tumor
immunotherapy (Ribas 2012, NEJM 366: 2517-2519; Watanabe et al
2012, Clin. Dev. Immunol. Volume 2012, Article ID: 269756; Wang et
al 2013, J. Viral Hep. 20: 27-39).
[0004] T-cell co-stimulatory and co-inhibitory molecules
(collectively named co-signaling molecules) play a crucial role in
regulating T-cell activation, subset differentiation, effector
function and survival (Chen et al 2013, Nature Rev. Immunol. 13:
227-242). Following recognition of cognate peptide-MHC complexes on
antigen-presenting cells by the T-cell receptor, co-signaling
receptors co-localize with T-cell receptors at the immune synapse,
where they synergize with TCR signaling to promote or inhibit
T-cell activation and function (Flies et al 2011, Yale J. Biol.
Med. 84: 409-421). The ultimate immune response is regulated by a
balance between co-stimulatory and co-inhibitory signals ("immune
checkpoints") (Pardoll 2012, Nature 12: 252-264). PD-1 functions as
one such `immune checkpoint` in mediating peripheral T-cell
tolerance and in avoiding autoimmunity. PD-1 binds to PD-L1 or
PD-L2 and inhibits T-cell activation. The ability of PD1 to inhibit
T-cell activation is exploited by chronic viral infections and
tumors to evade immune response. In chronic viral infections, PD-1
is highly expressed on virus-specific T-cells and these T-cells
become "exhausted" with loss of effector functions and
proliferative capacity (Freeman 2008, PNAS 105: 10275-10276). PD-L1
is expressed on a wide variety of tumors and studies on animal
models have shown that PD-L1 on tumors inhibits T-cell activation
and lysis of tumor cells and may lead to increased death of
tumor-specific T-cells. The PD-1: PD-L1 system also plays an
important role in induced T-regulatory (Treg) cell development and
in sustaining Treg function (Francisco et al 2010, Immunol. Rev.
236: 219-242).
[0005] Since PD-1 plays an important role in autoimmunity, tumor
immunity and infectious immunity, it is an ideal target for
immunotherapy. Blocking PD-1 with antagonists, including monoclonal
antibodies, has been studied in treatments of cancer and chronic
viral infections (Sheridan 2012, Nature Biotechnology 30:
729-730).
[0006] Monoclonal antibodies to PD-1 are known in the art and have
been described, for example, in US Patent/Publication Nos. U.S.
Pat. Nos. 8,008,449, 8,168,757, 20110008369, 20130017199,
20130022595, and in WO2006121168, WO20091154335, WO2012145493,
WO2013014668, WO2009101611, EP2262837, and EP2504028.
BRIEF SUMMARY OF THE INVENTION
[0007] The present invention provides antibodies and
antigen-binding fragments thereof that bind PD-1. The antibodies of
the present invention are useful, inter alia, for targeting T cells
expressing PD-1, and for modulating PD-1 activity. In certain
embodiments, the antibodies of the invention are useful for
inhibiting or neutralizing PD-1 activity and/or for stimulating T
cell activation, e.g., under circumstances where T cell-mediated
killing is beneficial or desirable. In alternate embodiments, the
antibodies enhance PD-1 binding and/or activity and may be used to
inhibit T-cell activation. The anti-PD-1 antibodies of the
invention, or antigen-binding portions thereof, may be included as
part of a multi-specific antigen-binding molecule, for example, to
modulate the immune response and/or to target the antibodies to a
specific cell type, such as a tumor cell, an autoimmune tissue cell
or a virally infected cell. The antibodies are useful in treating a
disease or disorder such as cancer, viral infection and autoimmune
disease.
[0008] The antibodies of the invention can be full-length (for
example, an IgG1 or IgG4 antibody) or may comprise only an
antigen-binding portion (for example, a Fab, F(ab').sub.2 or scFv
fragment), and may be modified to affect functionality, e.g., to
eliminate residual effector functions (Reddy et al., 2000, J.
Immunol. 164:1925-1933). In certain embodiments, the antibodies may
be bispecific.
[0009] In a first aspect, the present invention provides isolated
recombinant monoclonal antibodies or antigen-binding fragments
thereof that bind specifically to PD-1. In certain embodiments, the
antibodies are fully human. Exemplary anti-PD-1 antibodies of the
present invention are listed in Tables 1-3 herein. Table 1 sets
forth the amino acid sequence identifiers of the heavy chain
variable regions (HCVRs), light chain variable regions (LCVRs),
heavy chain complementarity determining regions (HCDR1, HCDR2 and
HCDR3), and light chain complementarity determining regions (LCDR1,
LCDR2 and LCDR3) of the exemplary anti-PD-1 antibodies. Table 2
sets forth the nucleic acid sequence identifiers of the HCVRs,
LCVRs, HCDR1, HCDR2 HCDR3, LCDR1, LCDR2 and LCDR3 of the exemplary
anti-PD-1 antibodies. Table 3 sets forth the amino acid sequence
identifiers of heavy chain and light chain sequences of exemplary
anti-PD-1 antibodies.
[0010] The present invention provides antibodies, or
antigen-binding fragments thereof, comprising an HCVR comprising an
amino acid sequence selected from any of the HCVR amino acid
sequences listed in Table 1, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0011] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising an LCVR comprising an
amino acid sequence selected from any of the LCVR amino acid
sequences listed in Table 1, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0012] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising an HCVR and an LCVR
amino acid sequence pair (HCVR/LCVR) comprising any of the HCVR
amino acid sequences listed in Table 1 paired with any of the LCVR
amino acid sequences listed in Table 1. According to certain
embodiments, the present invention provides antibodies, or
antigen-binding fragments thereof, comprising an HCVR/LCVR amino
acid sequence pair contained within any of the exemplary anti-PD-1
antibodies listed in Table 1. In certain embodiments, the HCVR/LCVR
amino acid sequence pair is selected from the group consisting of
SEQ ID NOs: 2/10, 18/26, 34/42, 50/58, 66/74, 82/90, 98/106,
114/122, 130/138, 146/154, 162/170, 178/186, 194/202, 210/202,
218/202, 226/202, 234/202, 242/202, 250/202, 258/202, 266/202,
274/202, 282/202, 290/202, 298/186, 306/186 and 314/186. In certain
embodiments, the HCVR/LCVR amino acid sequence pair is selected
from one of SEQ ID NOs: 130/138 (e.g., H2M7795N), 162/170 (e.g.,
H2M7798N), 234/202 (e.g., H4xH9048P), or 314/186 (e.g.,
H4xH9008P).
[0013] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a heavy chain CDR1
(HCDR1) comprising an amino acid sequence selected from any of the
HCDR1 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0014] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a heavy chain CDR2
(HCDR2) comprising an amino acid sequence selected from any of the
HCDR2 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0015] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a heavy chain CDR3
(HCDR3) comprising an amino acid sequence selected from any of the
HCDR3 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0016] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a light chain CDR1
(LCDR1) comprising an amino acid sequence selected from any of the
LCDR1 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0017] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a light chain CDR2
(LCDR2) comprising an amino acid sequence selected from any of the
LCDR2 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0018] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a light chain CDR3
(LCDR3) comprising an amino acid sequence selected from any of the
LCDR3 amino acid sequences listed in Table 1 or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity.
[0019] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising an HCDR3 and an LCDR3
amino acid sequence pair (HCDR3/LCDR3) comprising any of the HCDR3
amino acid sequences listed in Table 1 paired with any of the LCDR3
amino acid sequences listed in Table 1. According to certain
embodiments, the present invention provides antibodies, or
antigen-binding fragments thereof, comprising an HCDR3/LCDR3 amino
acid sequence pair contained within any of the exemplary anti-PD-1
antibodies listed in Table 1. In certain embodiments, the
HCDR3/LCDR3 amino acid sequence pair is selected from the group
consisting of SEQ ID NOs: 136/144 (e.g., H2M7795N), 168/176 (e.g.,
H2M7798N), 240/208 (e.g., H4xH9048P), and 320/192 (e.g.,
H4xH9008P).
[0020] The present invention provides antibodies, or
antigen-binding fragments thereof, comprising a heavy chain
comprising an amino acid sequence selected from any of the HC amino
acid sequences listed in Table 3, or a substantially similar
sequence thereof having at least 90%, at least 95%, at least 98% or
at least 99% sequence identity thereto.
[0021] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a light chain
comprising an amino acid sequence selected from any of the LC amino
acid sequences listed in Table 3, or a substantially similar
sequence thereof having at least 90%, at least 95%, at least 98% or
at least 99% sequence identity thereto.
[0022] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a HC and a LC amino
acid sequence pair (HC/LC) comprising any of the HC amino acid
sequences listed in Table 3 paired with any of the LC amino acid
sequences listed in Table 3. According to certain embodiments, the
present invention provides antibodies, or antigen-binding fragments
thereof, comprising an HC/LC amino acid sequence pair contained
within any of the exemplary anti-PD-1 antibodies listed in Table 3.
In certain embodiments, the HC/LC amino acid sequence pair is
selected from the group consisting of SEQ ID NOs: 330/331, 332/333,
334/335, and 336/337.
[0023] The present invention also provides antibodies, or
antigen-binding fragments thereof, comprising a set of six CDRs
HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3) contained within any of the
exemplary anti-PD-1 antibodies listed in Table 1. In certain
embodiments, the HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3 amino acid
sequence set is selected from the group consisting of SEQ ID NOs:
132-134-136-140-142-144 (e.g., H2M7795N); 164-166-168-172-174-176
(e.g., H2M7798N); 236-238-240-204-206-208 (e.g., H4xH9048P); and
316-318-320-188-190-192 (e.g., H4xH9008P).
[0024] In a related embodiment, the present invention provides
antibodies, or antigen-binding fragments thereof, comprising a set
of six CDRs HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3) contained within
an HCVR/LCVR amino acid sequence pair as defined by any of the
exemplary anti-PD-1 antibodies listed in Table 1. For example, the
present invention includes antibodies, or antigen-binding fragments
thereof, comprising the HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3 amino
acid sequences set contained within an HCVR/LCVR amino acid
sequence pair selected from the group consisting of SEQ ID NOs:
130/138 (e.g., H2M7795N); 162/170 (e.g., H2M7798N); 234/202 (e.g.,
H4xH9048P); and 314/186 (e.g., H4xH9008P). Methods and techniques
for identifying CDRs within HCVR and LCVR amino acid sequences are
well known in the art and can be used to identify CDRs within the
specified HCVR and/or LCVR amino acid sequences disclosed herein.
Exemplary conventions that can be used to identify the boundaries
of CDRs include, e.g., the Kabat definition, the Chothia
definition, and the AbM definition. In general terms, the Kabat
definition is based on sequence variability, the Chothia definition
is based on the location of the structural loop regions, and the
AbM definition is a compromise between the Kabat and Chothia
approaches. See, e.g., Kabat, "Sequences of Proteins of
Immunological Interest," National Institutes of Health, Bethesda,
Md. (1991); Al-Lazikani et al., J. Mol. Biol. 273:927-948 (1997);
and Martin et al., Proc. Natl. Acad. Sci. USA 86:9268-9272 (1989).
Public databases are also available for identifying CDR sequences
within an antibody.
[0025] The present invention includes anti-PD-1 antibodies having a
modified glycosylation pattern. In some embodiments, modification
to remove undesirable glycosylation sites may be useful, or an
antibody lacking a fucose moiety present on the oligosaccharide
chain, for example, to increase antibody dependent cellular
cytotoxicity (ADCC) function (see Shield et al. (2002) JBC
277:26733). In other applications, modification of galactosylation
can be made in order to modify complement dependent cytotoxicity
(CDC).
[0026] The present invention also provides for antibodies and
antigen-binding fragments thereof that compete for specific binding
to PD-1 with an antibody or antigen-binding fragment thereof
comprising the CDRs of a HCVR and the CDRs of a LCVR, wherein the
HCVR and LCVR each has an amino acid sequence selected from the
HCVR and LCVR sequences listed in Table 1.
[0027] The present invention also provides isolated antibodies and
antigen-binding fragments thereof that block PD-1 binding to PD-L1
or PD-L2. In some embodiments, the antibody or antigen-binding
fragment thereof that blocks PD-1 binding to PD-L1 may bind to the
same epitope on PD-1 as PD-L1 or may bind to a different epitope on
PD-1 as PD-L1.
[0028] In alternate embodiments, the present invention provides
antibodies and antigen-binding fragments thereof that stimulate
PD-1 binding to PD-L1. In certain embodiments, the present
invention provides isolated antibodies or antigen-binding fragments
thereof that bind PD-1, wherein the antibodies or antigen-binding
fragments thereof enhance PD-1 binding to PD-L1. In some
embodiments, the isolated antibodies or antigen-binding fragments
thereof comprise the CDRs of a HCVR, wherein the HCVR has an amino
acid sequence selected from the group consisting of SEQ ID NOs: 2,
98, and 250; and the CDRs of a LCVR, wherein the LCVR has an amino
acid sequence selected from the group consisting of SEQ ID NOs: 10,
106, and 202. In some embodiments, the isolated antibodies or
antigen-binding fragments thereof comprise an HCVR/LCVR amino acid
sequence pair selected from the group consisting of SEQ ID NOs:
2/10 (e.g., H1M7789N), 98/106 (e.g., H2M7791N), and 250/202 (e.g.,
H4H9068P2).
[0029] The present invention also provides antibodies and
antigen-binding fragments thereof that bind specifically to PD-1
from human or other species. In certain embodiments, the antibodies
may bind to human PD-1 and/or to cynomolgus PD-1.
[0030] The present invention also provides antibodies and
antigen-binding fragments thereof that cross-compete for binding to
PD-1 with a reference antibody or antigen-binding fragment thereof
comprising the CDRs of a HCVR and the CDRs of a LCVR, wherein the
HCVR and LCVR each has an amino acid sequence selected from the
HCVR and LCVR sequences listed in Table 1.
[0031] In one embodiment, the invention provides an isolated
antibody or antigen-binding fragment that has one or more of the
following characteristics: (a) blocks the binding of PD-1 to PD-L1
or PD-L2; (b) binds specifically to human PD-1 and/or cynomolgus
PD-1; (c) blocks PD-1-induced T-cell down regulation and rescues
T-cell signaling; (d) suppresses tumor growth and increases
survival in subjects with colon cancer; (e) inhibits T-cell
proliferation in a mixed lymphocyte reaction (MLR) assay; and (f)
increases IL-2 and/or interferon-gamma secretion in a MLR
assay.
[0032] In some embodiments, the antibody or antigen binding
fragment thereof may bind specifically to PD-1 in an agonist
manner, i.e., it may enhance or stimulate PD-1 binding and/or
activity; in other embodiments, the antibody may bind specifically
to PD-1 in an antagonist manner, i.e., it may block PD-1 from
binding to its ligand.
[0033] In certain embodiments, the antibodies or antigen-binding
fragments of the present invention are bispecific comprising a
first binding specificity to PD-1 and a second binding specificity
for a second target epitope. The second target epitope may be
another epitope on PD-1 or on a different protein. In certain
embodiments, the target epitope may be on a different cell
including a different T-cell, a B-cell, a tumor cell, an autoimmune
tissue cell or a virally infected cell.
[0034] In a second aspect, the present invention provides nucleic
acid molecules encoding anti-PD-1 antibodies or portions thereof.
For example, the present invention provides nucleic acid molecules
encoding any of the HCVR amino acid sequences listed in Table 1; in
certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the HCVR nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0035] The present invention also provides nucleic acid molecules
encoding any of the LCVR amino acid sequences listed in Table 1; in
certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the LCVR nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0036] The present invention also provides nucleic acid molecules
encoding any of the HCDR1 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the HCDR1 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0037] The present invention also provides nucleic acid molecules
encoding any of the HCDR2 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the HCDR2 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0038] The present invention also provides nucleic acid molecules
encoding any of the HCDR3 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the HCDR3 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0039] The present invention also provides nucleic acid molecules
encoding any of the LCDR1 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the LCDR1 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0040] The present invention also provides nucleic acid molecules
encoding any of the LCDR2 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the LCDR2 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0041] The present invention also provides nucleic acid molecules
encoding any of the LCDR3 amino acid sequences listed in Table 1;
in certain embodiments the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the LCDR3 nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto.
[0042] The present invention also provides nucleic acid molecules
encoding an HCVR, wherein the HCVR comprises a set of three CDRs
(i.e., HCDR1-HCDR2-HCDR3), wherein the HCDR1-HCDR2-HCDR3 amino acid
sequence set is as defined by any of the exemplary anti-PD-1
antibodies listed in Table 1.
[0043] The present invention also provides nucleic acid molecules
encoding an LCVR, wherein the LCVR comprises a set of three CDRs
(i.e., LCDR1-LCDR2-LCDR3), wherein the LCDR1-LCDR2-LCDR3 amino acid
sequence set is as defined by any of the exemplary anti-PD-1
antibodies listed in Table 1.
[0044] The present invention also provides nucleic acid molecules
encoding both an HCVR and an LCVR, wherein the HCVR comprises an
amino acid sequence of any of the HCVR amino acid sequences listed
in Table 1, and wherein the LCVR comprises an amino acid sequence
of any of the LCVR amino acid sequences listed in Table 1. In
certain embodiments, the nucleic acid molecule comprises a
polynucleotide sequence selected from any of the HCVR nucleic acid
sequences listed in Table 2, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity thereto, and a polynucleotide sequence
selected from any of the LCVR nucleic acid sequences listed in
Table 2, or a substantially similar sequence thereof having at
least 90%, at least 95%, at least 98% or at least 99% sequence
identity thereto. In certain embodiments according to this aspect
of the invention, the nucleic acid molecule encodes an HCVR and
LCVR, wherein the HCVR and LCVR are both derived from the same
anti-PD-1 antibody listed in Table 1.
[0045] The present invention provides nucleic acid molecules
encoding any of the heavy chain amino acid sequences listed in
Table 3. The present invention also provides nucleic acid molecules
encoding any of the light chain amino acid sequences listed in
Table 3.
[0046] The present invention also provides nucleic acid molecules
encoding both heavy chain (HC) and a light chain (LC), wherein the
HC comprises an amino acid sequence of any of the HC amino acid
sequences listed in Table 3, and wherein the LC comprises an amino
acid sequence of any of the LC amino acid sequences listed in Table
3.
[0047] In a related aspect, the present invention provides
recombinant expression vectors capable of expressing a polypeptide
comprising a heavy or light chain variable region of an anti-PD-1
antibody. For example, the present invention includes recombinant
expression vectors comprising any of the nucleic acid molecules
mentioned above, i.e., nucleic acid molecules encoding any of the
HCVR, LCVR, and/or CDR sequences as set forth in Table 1. The
present invention also provides recombinant expression vectors
capable of expressing a polypeptide comprising a heavy or light
chain of an anti-PD-1 antibody. For example, the present invention
includes recombinant expression vectors comprising any of the
nucleic acid molecules mentioned above, i.e., nucleic acid
molecules encoding any of the heavy chain or light chain sequences
as set forth in Table 3. Also included within the scope of the
present invention are host cells into which such vectors have been
introduced, as well as methods of producing the antibodies or
portions thereof by culturing the host cells under conditions
permitting production of the antibodies or antibody fragments, and
recovering the antibodies and antibody fragments so produced.
[0048] In a third aspect, the present invention provides
multi-specific antigen-binding molecules and antigen-binding
fragments thereof comprising a first antigen-binding specificity
that binds specifically to PD-1 and a second antigen-binding
specificity that binds specifically to an antigen selected from the
group consisting of a tumor cell-specific antigen, an autoimmune
tissue-specific antigen, an infected-cell-specific antigen, a
T-cell co-inhibitor, a T-cell receptor, a Fc receptor, PD-L1, and
PD-1. In certain embodiments, the first antigen-binding specificity
may comprise three CDRs derived from a HCVR with an amino acid
sequence selected from the HCVR sequences in Table 1 and three CDRs
derived from a LCVR with an amino acid sequence selected from the
LCVR sequences in Table 1. In one embodiment, the first
antigen-binding specificity may comprise the extracellular domain
of PD-L1. The second antigen-binding specificity may target an
antigen on the same cell as PD-1 or on a different cell of the same
tissue type or of a different tissue type. For example, the
multi-specific antigen-binding molecule may bind to a T-cell
wherein the first antigen-binding specificity may bind specifically
to PD-1 and the second antigen-binding specificity may bind to a
T-cell receptor on the T-cell. Alternatively, in another
embodiment, the first antigen-binding specificity may bind
specifically to PD-1 on a T-cell and the second antigen-binding
specificity may be targeted to an antigen/receptor on a B-cell or a
macrophage or antigen-presenting cell. In certain embodiments, the
second antigen-binding specificity may be directed to an antigen
associated with an autoimmune tissue. In one embodiment, the first
antigen-binding specificity may comprise an extracellular domain of
PD-L1 and the second antigen-binding specificity may bind to
another epitope on PD-1. In certain embodiments, the first
antigen-binding specificity binds to PD-1 with a lower affinity,
for example, with a K.sub.D more than 10.sup.-7 M, more than
10.sup.-6 M, more than 10.sup.-5 M, or more than 10.sup.-4 M.
[0049] In a fourth aspect, the invention provides a pharmaceutical
composition comprising a recombinant human antibody or fragment
thereof which specifically binds PD-1 and a pharmaceutically
acceptable carrier. In a related aspect, the invention features a
composition which is a combination of an anti-PD-1 antibody and a
second therapeutic agent. In one embodiment, the second therapeutic
agent is any agent that is advantageously combined with an
anti-PD-1 antibody. Exemplary agents that may be advantageously
combined with an anti-PD-1 antibody include, without limitation,
other agents that bind and/or modulate PD-1 signaling (including
other antibodies or antigen-binding fragments thereof, etc.) and/or
agents which do not directly bind PD-1 but nonetheless modulate
immune cell activation. Additional combination therapies and
co-formulations involving the anti-PD-1 antibodies of the present
invention are disclosed elsewhere herein.
[0050] In a fifth aspect, the invention provides methods to
modulate the immune response in a subject, the method comprising
administering a therapeutically effective amount of an anti-PD-1
antibody or antigen-binding fragment thereof of the invention to
the subject in need thereof. In certain embodiments, the invention
provides methods to enhance the immune response in a subject, the
methods comprising administering to the subject an effective amount
of an antibody or fragment thereof of the invention that binds PD-1
and blocks PD-1 binding to PD-L1. In one embodiment, the invention
provides a method to stimulate or enhance T-cell stimulation in a
subject. In one embodiment, the invention provides methods to
inhibit a T-regulatory (Treg) cell in a subject, the methods
comprising administering a therapeutically effective amount of a
blocking antibody or antigen-binding fragment thereof of the
invention to the subject in need thereof. In certain embodiments,
the subject in need thereof may suffer from a disease or disorder
such as cancer or viral infection. In alternate embodiments, the
invention provides for methods to inhibit or suppress T-cell
activation in a subject, the methods comprising administering a
therapeutically effective amount of an activating antibody or
fragment thereof of the invention to the subject in need thereof.
In one embodiment, the subject may suffer from an autoimmune
disease or disorder.
[0051] In a sixth aspect, the invention provides therapeutic
methods for treating a disease or disorder such as cancer,
autoimmune disease or viral infection in a subject using an
anti-PD-1 antibody or antigen-binding portion of an antibody of the
invention, wherein the therapeutic methods comprise administering a
therapeutically effective amount of a pharmaceutical composition
comprising an antibody or fragment of an antibody of the invention
to the subject in need thereof. The disorder treated is any disease
or condition which is improved, ameliorated, inhibited or prevented
by stimulation or inhibition of PD-1 activity or signaling. In
certain embodiments, the antibody or antigen-binding fragment
thereof the invention is administered in combination with a second
therapeutic agent to the subject in need thereof. The second
therapeutic agent may be selected from the group consisting of an
antibody to another T-cell co-inhibitor, an antibody to a tumor
cell antigen, an antibody to a T-cell receptor, an antibody to a Fc
receptor, an antibody to an epitope on a virally infected cell, an
antibody to an autoimmune tissue antigen, an antibody to PD-L1, a
cytotoxic agent, an anti-cancer drug, an anti-viral drug, an
anti-inflammatory drug (e.g., corticosteroids), chemotherapeutic
agent, radiation therapy, an immunosuppressant and any other drug
or therapy known in the art. In certain embodiments, the second
therapeutic agent may be an agent that helps to counteract or
reduce any possible side effect(s) associated with an antibody or
antigen-binding fragment thereof of the invention, if such side
effect(s) should occur.
[0052] In certain embodiments, the present invention provides
methods for suppressing tumor growth. In certain embodiments, the
present invention provides methods to enhance survival of cancer
patients. Examples of cancer include, but are not limited to,
primary and/or recurrent cancer, including brain cancer (e.g.,
glioblastoma multiforme), lung cancer (e.g., non-small cell lung
cancer), squamous cell carcinoma of head and neck, renal cell
carcinoma, melanoma, multiple myeloma, prostate cancer, and colon
cancer. The methods comprise administering a pharmaceutical
composition comprising a therapeutically effective amount of an
anti-PD-1 antibody of the present invention in combination with a
second therapeutic agent selected from the group consisting of a
vascular endothelial growth factor (VEGF) antagonist (e.g.,
aflibercept, bevacizumab), an angiopoietin-2 (Ang2) inhibitor
(e.g., an anti-Ang2 antibody such as nesvacumab), a lymphocyte
activation gene 3 (LAG-3) inhibitor, a cytotoxic T-lymphocyte
antigen 4 (CTLA-4) inhibitor (e.g., ipilimumab), a chemotherapeutic
agent, and radiation therapy. Additional examples of additional
therapies/therapeutic agents that can be used in combination with
an anti-PD-1 antibody of the invention for use in treating cancer
are described elsewhere herein.
[0053] The antibody or fragment thereof may be administered
subcutaneously, intravenously, intradermally, intraperitoneally,
orally, intramuscularly, or intracranially. The antibody or
fragment thereof may be administered at a dose of about 0.1 mg/kg
of body weight to about 100 mg/kg of body weight of the
subject.
[0054] The present invention also includes use of an anti-PD-1
antibody or antigen-binding fragment thereof of the invention in
the manufacture of a medicament for the treatment of a disease or
disorder that would benefit from the blockade or enhancement of
PD-1 binding and/or signaling.
[0055] Other embodiments will become apparent from a review of the
ensuing detailed description.
BRIEF DESCRIPTION OF THE FIGURES
[0056] FIG. 1 is a schematic of the luciferase-based PD-1 bioassay
described in Example 8 herein. Panel A: Inactive Jurkat cells;
Panel B: Jurkat cells are activated by T-cell receptor (TCR)
clustering through the CD3xCD20 bispecific antibody; Panel C: PD-1
activation attenuates response in activated Jurkat cells; Panel D:
Blocking PD-1 rescues the response in activated Jurkat cells.
[0057] FIG. 2 illustrates tumor growth and survival results for
mice implanted with Colon-26 tumor cells at Day 0 and treated with
the indicated combinations of molecules by injection at Days 3, 6,
10, 13 and 19 ("early-treatment tumor model"). The graph depicts
tumor volume (in mm.sup.3) for the different experimental groups at
various time points after implantation. Upward arrows along the
X-axis indicate the timing of treatment injections. "mIgG2a" is
IgG2 isotype control; "Fc" is human Fc control; "VEGF Trap" is
aflibercept; "anti-PD-1" is anti-mouse PD-1 clone RPMI-14;
"anti-PD-L1" is an anti-PD-L1 monoclonal antibody as described
elsewhere herein.
[0058] FIG. 3 illustrates tumor growth and survival results for
mice implanted with Colon-26 tumor cells at Day 0 and treated with
the indicated combinations of molecules by injection at Days 3, 6,
10, 13 and 19 ("early-treatment tumor model"). The graph shows the
tumor volume (in mm.sup.3) of individual mice in each experimental
group at Day 28 after implantation. "mIgG2a" is IgG2 isotype
control; "Fc" is human Fc control; "VEGF Trap" is aflibercept;
"anti-PD-1" is anti-mouse PD-1 clone RPMI-14; "anti-PD-L1" is an
anti-PD-L1 monoclonal antibody as described elsewhere herein.
DETAILED DESCRIPTION
[0059] Before the present methods are described, it is to be
understood that this invention is not limited to particular
methods, and experimental conditions described, as such methods and
conditions may vary. It is also to be understood that the
terminology used herein is for the purpose of describing particular
embodiments only, and is not intended to be limiting, since the
scope of the present invention will be limited only by the appended
claims.
[0060] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention belongs. Although
any methods and materials similar or equivalent to those described
herein can be used in the practice or testing of the present
invention, preferred methods and materials are now described. All
publications mentioned herein are incorporated herein by reference
in their entirety.
[0061] The term "PD-1" refers to the programmed death-1 protein, a
T-cell co-inhibitor, also known as CD279. The amino acid sequence
of full-length PD-1 is provided in GenBank as accession number
NP_005009.2 and is also referred to herein as SEQ ID NO: 327. The
term "PD-1" also includes protein variants of PD-1 having the amino
acid sequence of SEQ ID NOs: 321, 322, 323, or 324. The term "PD-1"
includes recombinant PD-1 or a fragment thereof. The term also
encompasses PD-1 or a fragment thereof coupled to, for example,
histidine tag, mouse or human Fc, or a signal sequence such as
ROR1. For example, the term includes sequences exemplified by SEQ
ID NOs: 323 or 324, comprising a mouse Fc (mIgG2a) or human Fc
(hIgG1) at the C-terminal, coupled to amino acid residues 25-170 of
full-length PD-1 with a C93S change. Protein variants as
exemplified by SEQ ID NO: 321 comprise a histidine tag at the
C-terminal, coupled to amino acid residues 25-170 of full length
PD-1. Unless specified as being from a non-human species, the term
"PD-1" means human PD-1.
[0062] PD-1 is a member of the CD28/CTLA-4/ICOS family of T-cell
co-inhibitors. PD-1 is a 288-amino acid protein with an
extracellular N-terminal domain which is IgV-like, a transmembrane
domain and an intracellular domain containing an immunoreceptor
tyrosine-based inhibitory (ITIM) motif and an immunoreceptor
tyrosine-based switch (ITSM) motif (Chattopadhyay et al 2009,
Immunol. Rev.). The PD-1 receptor has two ligands, PD-ligand-1
(PD-L1) and PD-L2.
[0063] The term "PD-L1" refers to the ligand of the PD-1 receptor
also known as CD274 and B7H1. The amino acid sequence of
full-length PD-L1 is provided in GenBank as accession number
NP_054862.1 and is also referred to herein as SEQ ID NO: 328. The
term also encompasses PD-L1 or a fragment thereof coupled to, for
example, histidine tag, mouse or human Fc, or a signal sequence
such as ROR1. For example, the term includes sequences exemplified
by SEQ ID NOs: 325 or 326, comprising a mouse Fc (mIgG2a) or human
Fc (hIgG1) at the C-terminal, coupled to amino acid residues 19-239
of full-length PD-L1. PD-L1 is a 290 amino acid protein with an
extracellular IgV-like domain, a transmembrane domain and a highly
conserved intracellular domain of approximately 30 amino acids.
PD-L1 is constitutively expressed on many cells such as antigen
presenting cells (e.g., dendritic cells, macrophages, and B-cells)
and on hematopoietic and non-hematopoietic cells (e.g., vascular
endothelial cells, pancreatic islets, and sites of immune
privilege). PD-L1 is also expressed on a wide variety of tumors,
virally-infected cells and autoimmune tissue, and is a component of
the immunosuppressive milieu (Ribas 2012, NEJM 366: 2517-2519).
[0064] As used herein, the term "T-cell co-inhibitor" refers to a
ligand and/or receptor which modulates the immune response via
T-cell activation or suppression. The term "T-cell co-inhibitor",
also known as T-cell co-signaling molecule, includes, but is not
limited to, lymphocyte activation gene 3 protein (LAG-3, also known
as CD223), cytotoxic T-lymphocyte antigen-4 (CTLA-4), B and T
lymphocyte attenuator (BTLA), CD-28, 2B4, LY108, T-cell
immunoglobulin and mucin 3(TIM3), T-cell immunoreceptor with
immunoglobulin and ITIM (TIGIT; also known as VSIG9), leucocyte
associated immunoglobulin-like receptor 1 (LAIR1; also known as
CD305), inducible T-cell costimulator (ICOS; also known as CD278),
V-domain Ig suppressor of T-cell activation (VISTA) and CD160.
[0065] As used herein, the term "Fc receptor" refers to the surface
receptor protein found on immune cells including B lymphocytes,
natural killer cells, macrophages, basophils, neutrophils, and mast
cells, which has a binding specificity for the Fc region of an
antibody. The term "Fc receptor" includes, but is not limited to, a
Fc.gamma. receptor [e.g., Fc.gamma.RI (CD64), Fc.gamma.RIIA (CD32),
Fc.gamma.RIIB (CD32), Fc.gamma.RIIIA (CD16a), and Fc.gamma.RIIIB
(CD16b)], Fc.alpha. receptor (e.g., Fc.alpha.RI or CD89) and Fc
receptor [e.g., Fc RI, and Fc RII (CD23)].
[0066] The term "antibody", as used herein, is intended to refer to
immunoglobulin molecules comprised of four polypeptide chains, two
heavy (H) chains and two light (L) chains inter-connected by
disulfide bonds (i.e., "full antibody molecules"), as well as
multimers thereof (e.g. IgM) or antigen-binding fragments thereof.
Each heavy chain is comprised of a heavy chain variable region
("HCVR" or "V.sub.H") and a heavy chain constant region (comprised
of domains C.sub.H1, C.sub.H2 and C.sub.H3). Each light chain is
comprised of a light chain variable region ("LCVR or "V.sub.L") and
a light chain constant region (C.sub.L). The V.sub.H and V.sub.L
regions can be further subdivided into regions of hypervariability,
termed complementarity determining regions (CDR), interspersed with
regions that are more conserved, termed framework regions (FR).
Each V.sub.H and V.sub.L is composed of three CDRs and four FRs,
arranged from amino-terminus to carboxy-terminus in the following
order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. In certain embodiments
of the invention, the FRs of the antibody (or antigen binding
fragment thereof) may be identical to the human germline sequences,
or may be naturally or artificially modified. An amino acid
consensus sequence may be defined based on a side-by-side analysis
of two or more CDRs.
[0067] Substitution of one or more CDR residues or omission of one
or more CDRs is also possible. Antibodies have been described in
the scientific literature in which one or two CDRs can be dispensed
with for binding. Padlan et al. (1995 FASEB J. 9:133-139) analyzed
the contact regions between antibodies and their antigens, based on
published crystal structures, and concluded that only about one
fifth to one third of CDR residues actually contact the antigen.
Padlan also found many antibodies in which one or two CDRs had no
amino acids in contact with an antigen (see also, Vajdos et al.
2002 J Mol Biol 320:415-428).
[0068] CDR residues not contacting antigen can be identified based
on previous studies (for example residues H60-H65 in CDRH2 are
often not required), from regions of Kabat CDRs lying outside
Chothia CDRs, by molecular modeling and/or empirically. If a CDR or
residue(s) thereof is omitted, it is usually substituted with an
amino acid occupying the corresponding position in another human
antibody sequence or a consensus of such sequences. Positions for
substitution within CDRs and amino acids to substitute can also be
selected empirically. Empirical substitutions can be conservative
or non-conservative substitutions.
[0069] The fully human anti-PD-1 monoclonal antibodies disclosed
herein may comprise one or more amino acid substitutions,
insertions and/or deletions in the framework and/or CDR regions of
the heavy and light chain variable domains as compared to the
corresponding germline sequences. Such mutations can be readily
ascertained by comparing the amino acid sequences disclosed herein
to germline sequences available from, for example, public antibody
sequence databases. The present invention includes antibodies, and
antigen-binding fragments thereof, which are derived from any of
the amino acid sequences disclosed herein, wherein one or more
amino acids within one or more framework and/or CDR regions are
mutated to the corresponding residue(s) of the germline sequence
from which the antibody was derived, or to the corresponding
residue(s) of another human germline sequence, or to a conservative
amino acid substitution of the corresponding germline residue(s)
(such sequence changes are referred to herein collectively as
"germline mutations"). A person of ordinary skill in the art,
starting with the heavy and light chain variable region sequences
disclosed herein, can easily produce numerous antibodies and
antigen-binding fragments which comprise one or more individual
germline mutations or combinations thereof. In certain embodiments,
all of the framework and/or CDR residues within the V.sub.H and/or
V.sub.L domains are mutated back to the residues found in the
original germline sequence from which the antibody was derived. In
other embodiments, only certain residues are mutated back to the
original germline sequence, e.g., only the mutated residues found
within the first 8 amino acids of FR1 or within the last 8 amino
acids of FR4, or only the mutated residues found within CDR1, CDR2
or CDR3. In other embodiments, one or more of the framework and/or
CDR residue(s) are mutated to the corresponding residue(s) of a
different germline sequence (i.e., a germline sequence that is
different from the germline sequence from which the antibody was
originally derived). Furthermore, the antibodies of the present
invention may contain any combination of two or more germline
mutations within the framework and/or CDR regions, e.g., wherein
certain individual residues are mutated to the corresponding
residue of a particular germline sequence while certain other
residues that differ from the original germline sequence are
maintained or are mutated to the corresponding residue of a
different germline sequence. Once obtained, antibodies and
antigen-binding fragments that contain one or more germline
mutations can be easily tested for one or more desired property
such as, improved binding specificity, increased binding affinity,
improved or enhanced antagonistic or agonistic biological
properties (as the case may be), reduced immunogenicity, etc.
Antibodies and antigen-binding fragments obtained in this general
manner are encompassed within the present invention.
[0070] The present invention also includes fully human anti-PD-1
monoclonal antibodies comprising variants of any of the HCVR, LCVR,
and/or CDR amino acid sequences disclosed herein having one or more
conservative substitutions. For example, the present invention
includes anti-PD-1 antibodies having HCVR, LCVR, and/or CDR amino
acid sequences with, e.g., 10 or fewer, 8 or fewer, 6 or fewer, 4
or fewer, etc. conservative amino acid substitutions relative to
any of the HCVR, LCVR, and/or CDR amino acid sequences disclosed
herein.
[0071] The term "human antibody", as used herein, is intended to
include antibodies having variable and constant regions derived
from human germline immunoglobulin sequences. The human mAbs of the
invention may include amino acid residues not encoded by human
germline immunoglobulin sequences (e.g., mutations introduced by
random or site-specific mutagenesis in vitro or by somatic mutation
in vivo), for example in the CDRs and in particular CDR3. However,
the term "human antibody", as used herein, is not intended to
include mAbs in which CDR sequences derived from the germline of
another mammalian species (e.g., mouse), have been grafted onto
human FR sequences. The term includes antibodies recombinantly
produced in a non-human mammal, or in cells of a non-human mammal.
The term is not intended to include antibodies isolated from or
generated in a human subject.
[0072] The term "recombinant", as used herein, refers to antibodies
or antigen-binding fragments thereof of the invention created,
expressed, isolated or obtained by technologies or methods known in
the art as recombinant DNA technology which include, e.g., DNA
splicing and transgenic expression. The term refers to antibodies
expressed in a non-human mammal (including transgenic non-human
mammals, e.g., transgenic mice), or a cell (e.g., CHO cells)
expression system or isolated from a recombinant combinatorial
human antibody library.
[0073] The term "multi-specific antigen-binding molecules", as used
herein refers to bispecific, tri-specific or multi-specific
antigen-binding molecules, and antigen-binding fragments thereof.
Multi-specific antigen-binding molecules may be specific for
different epitopes of one target polypeptide or may contain
antigen-binding domains specific for epitopes of more than one
target polypeptide. A multi-specific antigen-binding molecule can
be a single multifunctional polypeptide, or it can be a multimeric
complex of two or more polypeptides that are covalently or
non-covalently associated with one another. The term
"multi-specific antigen-binding molecules" includes antibodies of
the present invention that may be linked to or co-expressed with
another functional molecule, e.g., another peptide or protein. For
example, an antibody or fragment thereof can be functionally linked
(e.g., by chemical coupling, genetic fusion, non-covalent
association or otherwise) to one or more other molecular entities,
such as a protein or fragment thereof to produce a bi-specific or a
multi-specific antigen-binding molecule with a second binding
specificity. According to the present invention, the term
"multi-specific antigen-binding molecules" also includes
bi-specific, tri-specific or multi-specific antibodies or
antigen-binding fragments thereof. In certain embodiments, an
antibody of the present invention is functionally linked to another
antibody or antigen-binding fragment thereof to produce a
bispecific antibody with a second binding specificity. Bispecific
and multi-specific antibodies of the present invention are
described elsewhere herein.
[0074] The term "specifically binds," or "binds specifically to",
or the like, means that an antibody or antigen-binding fragment
thereof forms a complex with an antigen that is relatively stable
under physiologic conditions. Specific binding can be characterized
by an equilibrium dissociation constant of at least about
1.times.10.sup.-8 M or less (e.g., a smaller K.sub.D denotes a
tighter binding). Methods for determining whether two molecules
specifically bind are well known in the art and include, for
example, equilibrium dialysis, surface plasmon resonance, and the
like. As described herein, antibodies have been identified by
surface plasmon resonance, e.g., BIACORE.TM., which bind
specifically to PD-1. Moreover, multi-specific antibodies that bind
to one domain in PD-1 and one or more additional antigens or a
bi-specific that binds to two different regions of PD-1 are
nonetheless considered antibodies that "specifically bind", as used
herein.
[0075] The term "high affinity" antibody refers to those mAbs
having a binding affinity to PD-1, expressed as K.sub.D, of at
least 10.sup.-7 M; preferably 10.sup.-8 M; more preferably
10.sup.-9M, even more preferably 10.sup.-10 M, even more preferably
10.sup.-11 M, as measured by surface plasmon resonance, e.g.,
BIACORE.TM. or solution-affinity ELISA.
[0076] By the term "slow off rate", "Koff" or "kd" is meant an
antibody that dissociates from PD-1, with a rate constant of
1.times.10.sup.-3 s.sup.-1 or less, preferably 1.times.10.sup.-4
s.sup.-1 or less, as determined by surface plasmon resonance, e.g.,
BIACORE.TM..
[0077] The terms "antigen-binding portion" of an antibody,
"antigen-binding fragment" of an antibody, and the like, as used
herein, include any naturally occurring, enzymatically obtainable,
synthetic, or genetically engineered polypeptide or glycoprotein
that specifically binds an antigen to form a complex. The terms
"antigen-binding fragment" of an antibody, or "antibody fragment",
as used herein, refers to one or more fragments of an antibody that
retain the ability to bind to PD-1.
[0078] In specific embodiments, antibody or antibody fragments of
the invention may be conjugated to a moiety such a ligand or a
therapeutic moiety ("immunoconjugate"), such as an antibiotic, a
second anti-PD-1 antibody, or an antibody to another antigen such a
tumor-specific antigen, an autoimmune tissue antigen, a
virally-infected cell antigen, a Fc receptor, a T-cell receptor, or
a T-cell co-inhibitor, or an immunotoxin, or any other therapeutic
moiety useful for treating a disease or condition including cancer,
autoimmune disease or chronic viral infection.
[0079] An "isolated antibody", as used herein, is intended to refer
to an antibody that is substantially free of other antibodies (Abs)
having different antigenic specificities (e.g., an isolated
antibody that specifically binds PD-1, or a fragment thereof, is
substantially free of Abs that specifically bind antigens other
than PD-1.
[0080] A "blocking antibody" or a "neutralizing antibody", as used
herein (or an "antibody that neutralizes PD-1 activity" or
"antagonist antibody"), is intended to refer to an antibody whose
binding to PD-1 results in inhibition of at least one biological
activity of PD-1. For example, an antibody of the invention may
prevent or block PD-1 binding to PD-L1.
[0081] An "activating antibody" or an "enhancing antibody", as used
herein (or an "agonist antibody"), is intended to refer to an
antibody whose binding to PD-1 results in increasing or stimulating
at least one biological activity of PD-1. For example, an antibody
of the invention may increase PD-1 binding to PD-L1.
[0082] The term "surface plasmon resonance", as used herein, refers
to an optical phenomenon that allows for the analysis of real-time
biomolecular interactions by detection of alterations in protein
concentrations within a biosensor matrix, for example using the
BIACORE.TM. system (Pharmacia Biosensor AB, Uppsala, Sweden and
Piscataway, N.J.).
[0083] The term "K.sub.D", as used herein, is intended to refer to
the equilibrium dissociation constant of a particular
antibody-antigen interaction.
[0084] The term "epitope" refers to an antigenic determinant that
interacts with a specific antigen binding site in the variable
region of an antibody molecule known as a paratope. A single
antigen may have more than one epitope. Thus, different antibodies
may bind to different areas on an antigen and may have different
biological effects. The term "epitope" also refers to a site on an
antigen to which B and/or T cells respond. It also refers to a
region of an antigen that is bound by an antibody. Epitopes may be
defined as structural or functional. Functional epitopes are
generally a subset of the structural epitopes and have those
residues that directly contribute to the affinity of the
interaction. Epitopes may also be conformational, that is, composed
of non-linear amino acids. In certain embodiments, epitopes may
include determinants that are chemically active surface groupings
of molecules such as amino acids, sugar side chains, phosphoryl
groups, or sulfonyl groups, and, in certain embodiments, may have
specific three-dimensional structural characteristics, and/or
specific charge characteristics.
[0085] The term "substantial identity" or "substantially
identical," when referring to a nucleic acid or fragment thereof,
indicates that, when optimally aligned with appropriate nucleotide
insertions or deletions with another nucleic acid (or its
complementary strand), there is nucleotide sequence identity in at
least about 90%, and more preferably at least about 95%, 96%, 97%,
98% or 99% of the nucleotide bases, as measured by any well-known
algorithm of sequence identity, such as FASTA, BLAST or GAP, as
discussed below. A nucleic acid molecule having substantial
identity to a reference nucleic acid molecule may, in certain
instances, encode a polypeptide having the same or substantially
similar amino acid sequence as the polypeptide encoded by the
reference nucleic acid molecule.
[0086] As applied to polypeptides, the term "substantial
similarity" or "substantially similar" means that two peptide
sequences, when optimally aligned, such as by the programs GAP or
BESTFIT using default gap weights, share at least 90% sequence
identity, even more preferably at least 95%, 98% or 99% sequence
identity. Preferably, residue positions, which are not identical,
differ by conservative amino acid substitutions. A "conservative
amino acid substitution" is one in which an amino acid residue is
substituted by another amino acid residue having a side chain (R
group) with similar chemical properties (e.g., charge or
hydrophobicity). In general, a conservative amino acid substitution
will not substantially change the functional properties of a
protein. In cases where two or more amino acid sequences differ
from each other by conservative substitutions, the percent or
degree of similarity may be adjusted upwards to correct for the
conservative nature of the substitution. Means for making this
adjustment are well known to those of skill in the art. See, e.g.,
Pearson (1994) Methods Mol. Biol. 24: 307-331, which is herein
incorporated by reference. Examples of groups of amino acids that
have side chains with similar chemical properties include 1)
aliphatic side chains: glycine, alanine, valine, leucine and
isoleucine; 2) aliphatic-hydroxyl side chains: serine and
threonine; 3) amide-containing side chains: asparagine and
glutamine; 4) aromatic side chains: phenylalanine, tyrosine, and
tryptophan; 5) basic side chains: lysine, arginine, and histidine;
6) acidic side chains: aspartate and glutamate, and 7)
sulfur-containing side chains: cysteine and methionine. Preferred
conservative amino acids substitution groups are:
valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine,
alanine-valine, glutamate-aspartate, and asparagine-glutamine.
Alternatively, a conservative replacement is any change having a
positive value in the PAM250 log-likelihood matrix disclosed in
Gonnet et al. (1992) Science 256: 1443 45, herein incorporated by
reference. A "moderately conservative" replacement is any change
having a nonnegative value in the PAM250 log-likelihood matrix.
[0087] Sequence similarity for polypeptides is typically measured
using sequence analysis software. Protein analysis software matches
similar sequences using measures of similarity assigned to various
substitutions, deletions and other modifications, including
conservative amino acid substitutions. For instance, GCG software
contains programs such as GAP and BESTFIT which can be used with
default parameters to determine sequence homology or sequence
identity between closely related polypeptides, such as homologous
polypeptides from different species of organisms or between a wild
type protein and a mutein thereof. See, e.g., GCG Version 6.1.
Polypeptide sequences also can be compared using FASTA with default
or recommended parameters; a program in GCG Version 6.1. FASTA
(e.g., FASTA2 and FASTA3) provides alignments and percent sequence
identity of the regions of the best overlap between the query and
search sequences (Pearson (2000) supra). Another preferred
algorithm when comparing a sequence of the invention to a database
containing a large number of sequences from different organisms is
the computer program BLAST, especially BLASTP or TBLASTN, using
default parameters. See, e.g., Altschul et al. (1990) J. Mol. Biol.
215: 403-410 and (1997) Nucleic Acids Res. 25:3389-3402, each of
which is herein incorporated by reference.
[0088] By the phrase "therapeutically effective amount" is meant an
amount that produces the desired effect for which it is
administered. The exact amount will depend on the purpose of the
treatment, and will be ascertainable by one skilled in the art
using known techniques (see, for example, Lloyd (1999) The Art,
Science and Technology of Pharmaceutical Compounding).
[0089] As used herein, the term "subject" refers to an animal,
preferably a mammal, in need of amelioration, prevention and/or
treatment of a disease or disorder such as chronic viral infection,
cancer or autoimmune disease.
[0090] As used herein, "anti-cancer drug" means any agent useful to
treat cancer including, but not limited to, cytotoxins and agents
such as antimetabolites, alkylating agents, anthracyclines,
antibiotics, antimitotic agents, procarbazine, hydroxyurea,
asparaginase, corticosteroids, mytotane (O,P'-(DDD)), biologics
(e.g., antibodies and interferons) and radioactive agents. As used
herein, "a cytotoxin or cytotoxic agent", also refers to a
chemotherapeutic agent and means any agent that is detrimental to
cells. Examples include Taxol.RTM. (paclitaxel), temozolamide,
cytochalasin B, gramicidin D, ethidium bromide, emetine, cisplatin,
mitomycin, etoposide, tenoposide, vincristine, vinbiastine,
coichicin, doxorubicin, daunorubicin, dihydroxy anthracin dione,
mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone,
glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and
puromycin and analogs or homologs thereof.
[0091] As used herein, the term "anti-viral drug" refers to any
drug or therapy used to treat, prevent, or ameliorate a viral
infection in a host subject. The term "anti-viral drug" includes,
but is not limited to zidovudine, lamivudine, abacavir, ribavirin,
lopinavir, efavirenz, cobicistat, tenofovir, rilpivirine,
analgesics and corticosteroids. In the context of the present
invention, the viral infections include long-term or chronic
infections caused by viruses including, but not limited to, human
immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C
virus (HCV), human papilloma virus (HPV), lymphocytic
choriomeningitis virus (LCMV), and simian immunodeficiency virus
(SIV).
[0092] The antibodies and antigen-binding fragments of the present
invention specifically bind to PD-1 and modulate the interaction of
PD-1 with PD-L1. The anti-PD-1 antibodies may bind to PD-1 with
high affinity or with low affinity. In certain embodiments, the
antibodies of the present invention may be blocking antibodies
wherein the antibodies may bind to PD-1 and block the interaction
of PD-1 with PD-L1. In some embodiments, the blocking antibodies of
the invention may block the binding of PD-1 to PD-L1 and/or
stimulate or enhance T-cell activation. In some embodiments, the
blocking antibodies may be useful for stimulating or enhancing the
immune response and/or for treating a subject suffering from
cancer, or a chronic viral infection. The antibodies when
administered to a subject in need thereof may reduce the chronic
infection by a virus such as HIV, LCMV or HBV in the subject. They
may be used to inhibit the growth of tumor cells in a subject. They
may be used alone or as adjunct therapy with other therapeutic
moieties or modalities known in the art for treating cancer, or
viral infection.
[0093] In other embodiments, the antibodies of the present
invention may be activating antibodies, wherein the antibodies may
bind to PD-1 and enhance the interaction of PD-1 and PD-L1. In some
embodiments, the activating antibodies may enhance binding of PD-1
to PD-L1 and/or inhibit or suppress T-cell activation. The
activating antibodies of the present invention may be useful for
inhibiting the immune response in a subject and/or for treating
autoimmune disease.
[0094] In certain embodiments, the anti-PD-1 antibodies may be
multi-specific antigen-binding molecules, wherein they comprise a
first binding specificity to PD-1 and a second binding specificity
to an antigen selected from the group consisting of another T-cell
co-inhibitor, an autoimmune tissue antigen, T-cell receptor, Fc
receptor, T-cell receptor, PD-L1, and a different epitope of
PD-1.
[0095] In certain embodiments, the antibodies of the invention are
obtained from mice immunized with a primary immunogen, such as a
full length PD-1 [See GenBank accession number NP_005009.2 (SEQ ID
NO: 327)] or with a recombinant form of PD-1 or modified human PD-1
fragments (SEQ ID NOs: 321, 323, or 324) or with modified
cynomolgus PD-1 fragments (SEQ ID NO: 322), followed by
immunization with a secondary immunogen, or with an immunogenically
active fragment of PD-1.
[0096] The immunogen may be a biologically active and/or
immunogenic fragment of PD-1 or DNA encoding the active fragment
thereof. The fragment may be derived from the N-terminal or
C-terminal domain of PD-1. In certain embodiments of the invention,
the immunogen is a fragment of PD-1 that ranges from amino acid
residues 25-170 of SEQ ID NO: 327 with a C93S change.
[0097] The peptides may be modified to include addition or
substitution of certain residues for tagging or for purposes of
conjugation to carrier molecules, such as, KLH. For example, a
cysteine may be added at either the N terminal or C terminal end of
a peptide, or a linker sequence may be added to prepare the peptide
for conjugation to, for example, KLH for immunization.
[0098] The full-length amino acid sequence of full length human
PD-1 is shown as SEQ ID NO: 327.
[0099] In certain embodiments, antibodies that bind specifically to
PD-1 may be prepared using fragments of the above-noted regions, or
peptides that extend beyond the designated regions by about 5 to
about 20 amino acid residues from either, or both, the N or C
terminal ends of the regions described herein. In certain
embodiments, any combination of the above-noted regions or
fragments thereof may be used in the preparation of PD-1 specific
antibodies. In certain embodiments, any one or more of the
above-noted regions of PD-1, or fragments thereof may be used for
preparing monospecific, bispecific, or multispecific
antibodies.
[0100] Certain anti-PD-1 antibodies of the present invention are
able to bind to and neutralize the activity of PD-1, as determined
by in vitro or in vivo assays. The ability of the antibodies of the
invention to bind to and neutralize the activity of PD-1 may be
measured using any standard method known to those skilled in the
art, including binding assays, or activity assays, as described
herein.
[0101] Non-limiting, exemplary in vitro assays for measuring
binding activity are illustrated in Examples herein. In Example 3,
the binding affinities and kinetic constants of human anti-PD-1
antibodies for human PD-1 and cynomolgus PD-1 were determined by
surface plasmon resonance and the measurements were conducted on a
Biacore 4000 or T200 instrument. In Examples 4 and 5, blocking
assays were used to determine the ability of the anti-PD-1
antibodies to block PD-L1-binding ability of PD-1 in vitro. In
Example 6, blocking assays were used to determine cross-competition
between anti-PD-1 antibodies. Example 7 describes the binding of
the antibodies to cells overexpressing PD-1. In Example 8, a
luciferase assay was used to determine the ability of anti-PD-1
antibodies to antagonize PD-1/PD-L1 signaling in T-cells.
[0102] In certain embodiments, the antibodies of the present
invention are able to enhance or stimulate T-cell activation in
vitro and in a subject with cancer or in a subject infected with a
virus such as LCMV. In certain embodiments, the antibodies of the
present invention are used in combination with a second therapeutic
agent, such as an antibody to a second T-cell co-inhibitor, to
enhance the immune response and inhibit tumor growth in a
subject.
[0103] The antibodies specific for PD-1 may contain no additional
labels or moieties, or they may contain an N-terminal or C-terminal
label or moiety. In one embodiment, the label or moiety is biotin.
In a binding assay, the location of a label (if any) may determine
the orientation of the peptide relative to the surface upon which
the peptide is bound. For example, if a surface is coated with
avidin, a peptide containing an N-terminal biotin will be oriented
such that the C-terminal portion of the peptide will be distal to
the surface. In one embodiment, the label may be a radionuclide, a
fluorescent dye or a MRI-detectable label. In certain embodiments,
such labeled antibodies may be used in diagnostic assays including
imaging assays.
Antigen-Binding Fragments of Antibodies
[0104] Unless specifically indicated otherwise, the term
"antibody," as used herein, shall be understood to encompass
antibody molecules comprising two immunoglobulin heavy chains and
two immunoglobulin light chains (i.e., "full antibody molecules")
as well as antigen-binding fragments thereof. The terms
"antigen-binding portion" of an antibody, "antigen-binding
fragment" of an antibody, and the like, as used herein, include any
naturally occurring, enzymatically obtainable, synthetic, or
genetically engineered polypeptide or glycoprotein that
specifically binds an antigen to form a complex. The terms
"antigen-binding fragment" of an antibody, or "antibody fragment",
as used herein, refers to one or more fragments of an antibody that
retain the ability to specifically bind to PD-1. An antibody
fragment may include a Fab fragment, a F(ab').sub.2 fragment, a Fv
fragment, a dAb fragment, a fragment containing a CDR, or an
isolated CDR. In certain embodiments, the term "antigen-binding
fragment" refers to a polypeptide fragment of a multi-specific
antigen-binding molecule. In such embodiments, the term"
antigen-binding fragment" includes, e.g., an extracellular domain
of PD-L1 which binds specifically to PD-1. Antigen-binding
fragments of an antibody may be derived, e.g., from full antibody
molecules using any suitable standard techniques such as
proteolytic digestion or recombinant genetic engineering techniques
involving the manipulation and expression of DNA encoding antibody
variable and (optionally) constant domains. Such DNA is known
and/or is readily available from, e.g., commercial sources, DNA
libraries (including, e.g., phage-antibody libraries), or can be
synthesized. The DNA may be sequenced and manipulated chemically or
by using molecular biology techniques, for example, to arrange one
or more variable and/or constant domains into a suitable
configuration, or to introduce codons, create cysteine residues,
modify, add or delete amino acids, etc.
[0105] Non-limiting examples of antigen-binding fragments include:
(i) Fab fragments; (ii) F(ab')2 fragments; (iii) Fd fragments; (iv)
Fv fragments; (v) single-chain Fv (scFv) molecules; (vi) dAb
fragments; and (vii) minimal recognition units consisting of the
amino acid residues that mimic the hypervariable region of an
antibody (e.g., an isolated complementarity determining region
(CDR) such as a CDR3 peptide), or a constrained FR3-CDR3-FR4
peptide. Other engineered molecules, such as domain-specific
antibodies, single domain antibodies, domain-deleted antibodies,
chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies,
tetrabodies, minibodies, nanobodies (e.g. monovalent nanobodies,
bivalent nanobodies, etc.), small modular immunopharmaceuticals
(SMIPs), and shark variable IgNAR domains, are also encompassed
within the expression "antigen-binding fragment," as used
herein.
[0106] An antigen-binding fragment of an antibody will typically
comprise at least one variable domain. The variable domain may be
of any size or amino acid composition and will generally comprise
at least one CDR, which is adjacent to or in frame with one or more
framework sequences. In antigen-binding fragments having a V.sub.H
domain associated with a V.sub.L domain, the V.sub.H and V.sub.L
domains may be situated relative to one another in any suitable
arrangement. For example, the variable region may be dimeric and
contain V.sub.H-V.sub.H, V.sub.H-V.sub.L or V.sub.L-V.sub.L dimers.
Alternatively, the antigen-binding fragment of an antibody may
contain a monomeric V.sub.H or V.sub.L domain.
[0107] In certain embodiments, an antigen-binding fragment of an
antibody may contain at least one variable domain covalently linked
to at least one constant domain. Non-limiting, exemplary
configurations of variable and constant domains that may be found
within an antigen-binding fragment of an antibody of the present
invention include: (i) V.sub.H-C.sub.H1; (ii) V.sub.H-C.sub.H2;
(iii) V.sub.H-C.sub.H3; (iv) V.sub.H-C.sub.H1-C.sub.H2; (v)
V.sub.H-C.sub.H1-C.sub.H2-C.sub.H3; (vi) V.sub.H-C.sub.H2-C.sub.H3;
(vii) V.sub.H-C.sub.L; (viii) V.sub.L-C.sub.H1; (ix) V.sub.L-
C.sub.H2; (x) V.sub.L-C.sub.H3; (xi) V.sub.L-C.sub.H1-C.sub.H2;
(xii) V.sub.L-C.sub.H1-C.sub.H2-C.sub.H3; (xiii)
V.sub.L-C.sub.H2-C.sub.H3; and (xiv) V.sub.L-C.sub.L. In any
configuration of variable and constant domains, including any of
the exemplary configurations listed above, the variable and
constant domains may be either directly linked to one another or
may be linked by a full or partial hinge or linker region. A hinge
region may consist of at least 2 (e.g., 5, 10, 15, 20, 40, 60 or
more) amino acids, which result in a flexible or semi-flexible
linkage between adjacent variable and/or constant domains in a
single polypeptide molecule. Moreover, an antigen-binding fragment
of an antibody of the present invention may comprise a homo-dimer
or hetero-dimer (or other multimer) of any of the variable and
constant domain configurations listed above in non-covalent
association with one another and/or with one or more monomeric
V.sub.H or V.sub.L domain (e.g., by disulfide bond(s)).
[0108] As with full antibody molecules, antigen-binding fragments
may be mono-specific or multi-specific (e.g., bi-specific). A
multi-specific antigen-binding fragment of an antibody will
typically comprise at least two different variable domains, wherein
each variable domain is capable of specifically binding to a
separate antigen or to a different epitope on the same antigen. Any
multi-specific antibody format, including the exemplary bi-specific
antibody formats disclosed herein, may be adapted for use in the
context of an antigen-binding fragment of an antibody of the
present invention using routine techniques available in the
art.
Preparation of Human Antibodies
[0109] Methods for generating human antibodies in transgenic mice
are known in the art. Any such known methods can be used in the
context of the present invention to make human antibodies that
specifically bind to PD-1.
[0110] An immunogen comprising any one of the following can be used
to generate antibodies to PD-1. In certain embodiments, the
antibodies of the invention are obtained from mice immunized with a
full length, native PD-1 (See GenBank accession number NP_005009.2)
(SEQ ID NO: 327), or with a recombinant PD-1 peptide.
Alternatively, PD-1 or a fragment thereof may be produced using
standard biochemical techniques and modified (SEQ ID NOS: 321-324)
and used as immunogen.
[0111] In certain embodiments, the immunogen may be a peptide from
the N terminal or C terminal end of PD-1. In one embodiment, the
immunogen is the extracellular domain or the IgV-like domain of
PD-1. In certain embodiments of the invention, the immunogen is a
fragment of PD-1 that ranges from about amino acid residues 25-170
of SEQ ID NO: 327 with a C93S change.
[0112] In some embodiments, the immunogen may be a recombinant PD-1
peptide expressed in E. coli or in any other eukaryotic or
mammalian cells such as Chinese hamster ovary (CHO) cells.
[0113] In certain embodiments, antibodies that bind specifically to
PD-1 may be prepared using fragments of the above-noted regions, or
peptides that extend beyond the designated regions by about 5 to
about 20 amino acid residues from either, or both, the N or C
terminal ends of the regions described herein. In certain
embodiments, any combination of the above-noted regions or
fragments thereof may be used in the preparation of PD-1 specific
antibodies.
[0114] Using VELOCIMMUNE.RTM. technology (see, for example, U.S.
Pat. No. 6,596,541, Regeneron Pharmaceuticals, VELOCIMMUNE.RTM.) or
any other known method for generating monoclonal antibodies, high
affinity chimeric antibodies to PD-1 are initially isolated having
a human variable region and a mouse constant region. The
VELOCIMMUNE.RTM. technology involves generation of a transgenic
mouse having a genome comprising human heavy and light chain
variable regions operably linked to endogenous mouse constant
region loci such that the mouse produces an antibody comprising a
human variable region and a mouse constant region in response to
antigenic stimulation. The DNA encoding the variable regions of the
heavy and light chains of the antibody are isolated and operably
linked to DNA encoding the human heavy and light chain constant
regions. The DNA is then expressed in a cell capable of expressing
the fully human antibody.
Bioequivalents
[0115] The anti-PD-1 antibodies and antibody fragments of the
present invention encompass proteins having amino acid sequences
that vary from those of the described antibodies, but that retain
the ability to bind PD-1. Such variant antibodies and antibody
fragments comprise one or more additions, deletions, or
substitutions of amino acids when compared to parent sequence, but
exhibit biological activity that is essentially equivalent to that
of the described antibodies. Likewise, the antibody-encoding DNA
sequences of the present invention encompass sequences that
comprise one or more additions, deletions, or substitutions of
nucleotides when compared to the disclosed sequence, but that
encode an antibody or antibody fragment that is essentially
bioequivalent to an antibody or antibody fragment of the
invention.
[0116] Two antigen-binding proteins, or antibodies, are considered
bioequivalent if, for example, they are pharmaceutical equivalents
or pharmaceutical alternatives whose rate and extent of absorption
do not show a significant difference when administered at the same
molar dose under similar experimental conditions, either single
dose or multiple doses. Some antibodies will be considered
equivalents or pharmaceutical alternatives if they are equivalent
in the extent of their absorption but not in their rate of
absorption and yet may be considered bioequivalent because such
differences in the rate of absorption are intentional and are
reflected in the labeling, are not essential to the attainment of
effective body drug concentrations on, e.g., chronic use, and are
considered medically insignificant for the particular drug product
studied.
[0117] In one embodiment, two antigen-binding proteins are
bioequivalent if there are no clinically meaningful differences in
their safety, purity, or potency.
[0118] In one embodiment, two antigen-binding proteins are
bioequivalent if a patient can be switched one or more times
between the reference product and the biological product without an
expected increase in the risk of adverse effects, including a
clinically significant change in immunogenicity, or diminished
effectiveness, as compared to continued therapy without such
switching.
[0119] In one embodiment, two antigen-binding proteins are
bioequivalent if they both act by a common mechanism or mechanisms
of action for the condition or conditions of use, to the extent
that such mechanisms are known.
[0120] Bioequivalence may be demonstrated by in vivo and/or in
vitro methods. Bioequivalence measures include, e.g., (a) an in
vivo test in humans or other mammals, in which the concentration of
the antibody or its metabolites is measured in blood, plasma,
serum, or other biological fluid as a function of time; (b) an in
vitro test that has been correlated with and is reasonably
predictive of human in vivo bioavailability data; (c) an in vivo
test in humans or other mammals in which the appropriate acute
pharmacological effect of the antibody (or its target) is measured
as a function of time; and (d) in a well-controlled clinical trial
that establishes safety, efficacy, or bioavailability or
bioequivalence of an antibody.
[0121] Bioequivalent variants of the antibodies of the invention
may be constructed by, for example, making various substitutions of
residues or sequences or deleting terminal or internal residues or
sequences not needed for biological activity. For example, cysteine
residues not essential for biological activity can be deleted or
replaced with other amino acids to prevent formation of unnecessary
or incorrect intramolecular disulfide bridges upon renaturation. In
other contexts, bioequivalent antibodies may include antibody
variants comprising amino acid changes, which modify the
glycosylation characteristics of the antibodies, e.g., mutations
that eliminate or remove glycosylation.
Anti-PD-1 Antibodies Comprising Fc Variants
[0122] According to certain embodiments of the present invention,
anti-PD-1 antibodies are provided comprising an Fc domain
comprising one or more mutations which enhance or diminish antibody
binding to the FcRn receptor, e.g., at acidic pH as compared to
neutral pH. For example, the present invention includes anti-PD-1
antibodies comprising a mutation in the C.sub.H2 or a C.sub.H3
region of the Fc domain, wherein the mutation(s) increases the
affinity of the Fc domain to FcRn in an acidic environment (e.g.,
in an endosome where pH ranges from about 5.5 to about 6.0). Such
mutations may result in an increase in serum half-life of the
antibody when administered to an animal. Non-limiting examples of
such Fc modifications include, e.g., a modification at position 250
(e.g., E or Q); 250 and 428 (e.g., L or F); 252 (e.g., L/Y/F/W or
T), 254 (e.g., S or T), and 256 (e.g., S/R/Q/E/D or T); or a
modification at position 428 and/or 433 (e.g., H/L/R/S/P/Q or K)
and/or 434 (e.g., A, W, H, F or Y [N434A, N434W, N434H, N434F or
N434Y]); or a modification at position 250 and/or 428; or a
modification at position 307 or 308 (e.g., 308F, V308F), and 434.
In one embodiment, the modification comprises a 428L (e.g., M428L)
and 434S (e.g., N434S) modification; a 428L, 259I (e.g., V259I),
and 308F (e.g., V308F) modification; a 433K (e.g., H433K) and a 434
(e.g., 434Y) modification; a 252, 254, and 256 (e.g., 252Y, 254T,
and 256E) modification; a 250Q and 428L modification (e.g., T250Q
and M428L); and a 307 and/or 308 modification (e.g., 308F or 308P).
In yet another embodiment, the modification comprises a 265A (e.g.,
D265A) and/or a 297A (e.g., N297A) modification.
[0123] For example, the present invention includes anti-PD-1
antibodies comprising an Fc domain comprising one or more pairs or
groups of mutations selected from the group consisting of: 250Q and
248L (e.g., T250Q and M248L); 252Y, 254T and 256E (e.g., M252Y,
S254T and T256E); 428L and 434S (e.g., M428L and N434S); 257I and
311I (e.g., P257I and Q311I); 257I and 434H (e.g., P257I and
N434H); 376V and 434H (e.g., D376V and N434H); 307A, 380A and 434A
(e.g., T307A, E380A and N434A); and 433K and 434F (e.g., H433K and
N434F). In one embodiment, the present invention includes anti-PD-1
antibodies comprising an Fc domain comprising a S108P mutation in
the hinge region of IgG4 to promote dimer stabilization. All
possible combinations of the foregoing Fc domain mutations, and
other mutations within the antibody variable domains disclosed
herein, are contemplated within the scope of the present
invention.
[0124] The present invention also includes anti-PD-1 antibodies
comprising a chimeric heavy chain constant (C.sub.H) region,
wherein the chimeric C.sub.H region comprises segments derived from
the C.sub.H regions of more than one immunoglobulin isotype. For
example, the antibodies of the invention may comprise a chimeric
C.sub.H region comprising part or all of a C.sub.H2 domain derived
from a human IgG1, human IgG2 or human IgG4 molecule, combined with
part or all of a C.sub.H3 domain derived from a human IgG1, human
IgG2 or human IgG4 molecule. According to certain embodiments, the
antibodies of the invention comprise a chimeric C.sub.H region
having a chimeric hinge region. For example, a chimeric hinge may
comprise an "upper hinge" amino acid sequence (amino acid residues
from positions 216 to 227 according to EU numbering) derived from a
human IgG1, a human IgG2 or a human IgG4 hinge region, combined
with a "lower hinge" sequence (amino acid residues from positions
228 to 236 according to EU numbering) derived from a human IgG1, a
human IgG2 or a human IgG4 hinge region. According to certain
embodiments, the chimeric hinge region comprises amino acid
residues derived from a human IgG1 or a human IgG4 upper hinge and
amino acid residues derived from a human IgG2 lower hinge. An
antibody comprising a chimeric C.sub.H region as described herein
may, in certain embodiments, exhibit modified Fc effector functions
without adversely affecting the therapeutic or pharmacokinetic
properties of the antibody. (See, e.g., U.S. Ser. No. 14/170,166,
filed Jan. 31, 2014, the disclosure of which is hereby incorporated
by reference in its entirety).
Biological Characteristics of the Antibodies
[0125] In general, the antibodies of the present invention function
by binding to PD-1. The present invention includes anti-PD-1
antibodies and antigen-binding fragments thereof that bind soluble
monomeric or dimeric PD-1 molecules with high affinity. For
example, the present invention includes antibodies and
antigen-binding fragments of antibodies that bind monomeric PD-1
(e.g., at 25.degree. C. or at 37.degree. C.) with a K.sub.D of less
than about 50 nM as measured by surface plasmon resonance, e.g.,
using the assay format as defined in Example 3 herein. In certain
embodiments, the antibodies or antigen-binding fragments thereof
bind monomeric PD-1 with a K.sub.D of less than about 40 nM, less
than about 30 nM, less than about 20 nM, less than about 10 nM less
than about 5 nM, less than about 2 nM or less than about 1 nM, as
measured by surface plasmon resonance, e.g., using the assay format
as defined in Example 3 herein, or a substantially similar
assay.
[0126] The present invention also includes antibodies and
antigen-binding fragments thereof that bind dimeric PD-1 (e.g., at
25.degree. C. or at 37.degree. C.) with a K.sub.D of less than
about 400 pM as measured by surface plasmon resonance, e.g., using
the assay format as defined in Example 3 herein. In certain
embodiments, the antibodies or antigen-binding fragments thereof
bind dimeric PD-1 with a K.sub.D of less than about 300 pM, less
than about 250 pM, less than about 200 pM, less than about 100 pM,
or less than about 50 pM, as measured by surface plasmon resonance,
e.g., using the assay format as defined in Example 3 herein, or a
substantially similar assay.
[0127] The present invention also includes antibodies or
antigen-binding fragments thereof that bind cynomolgus (Macaca
fascicularis) PD-1 (e.g., at 25.degree. C. or at 37.degree. C.)
with a K.sub.D of less than about 35 nM as measured by surface
plasmon resonance, e.g., using the assay format as defined in
Example 3 herein. In certain embodiments, the antibodies or
antigen-binding fragments thereof bind cynomolgus PD-1 with a
K.sub.D of less than about 30 nM, less than about 20 nM, less than
about 15 nM, less than about 10 nM, or less than about 5 nM, as
measured by surface plasmon resonance, e.g., using the assay format
as defined in Example 3 herein, or a substantially similar
assay.
[0128] The present invention also includes antibodies and
antigen-binding fragments thereof that bind PD-1 with a
dissociative half-life (t1/2) of greater than about 1.1 minutes as
measured by surface plasmon resonance at 25.degree. C. or
37.degree. C., e.g., using an assay format as defined in Example 3
herein, or a substantially similar assay. In certain embodiments,
the antibodies or antigen-binding fragments of the present
invention bind PD-1 with a t1/2 of greater than about 5 minutes,
greater than about 10 minutes, greater than about 30 minutes,
greater than about 50 minutes, greater than about 60 minutes,
greater than about 70 minutes, greater than about 80 minutes,
greater than about 90 minutes, greater than about 100 minutes,
greater than about 200 minutes, greater than about 300 minutes,
greater than about 400 minutes, greater than about 500 minutes,
greater than about 600 minutes, greater than about 700 minutes,
greater than about 800 minutes, greater than about 900 minutes,
greater than about 1000 minutes, or greater than about 1200
minutes, as measured by surface plasmon resonance at 25.degree. C.
or 37.degree. C., e.g., using an assay format as defined in Example
3 herein (e.g., mAb-capture or antigen-capture format), or a
substantially similar assay.
[0129] The present invention also includes antibodies or
antigen-binding fragments thereof that block PD-1 binding to PD-L1
with an IC50 of less than about 3 nM as determined using a
ELISA-based immunoassay assay, e.g., as shown in Example 4, or a
substantially similar assay. The present invention also includes
antibodies and antigen-binding fragments thereof that bind to PD-1
and enhance the binding of PD-1 to PD-L1.
[0130] In some embodiments, the antibodies of the present invention
may bind to the extracellular domain of PD-1 or to a fragment of
the domain. In some embodiments, the antibodies of the present
invention may bind to more than one domain (cross-reactive
antibodies). In certain embodiments, the antibodies of the present
invention may bind to an epitope located in the extracellular
domain comprising amino acid residues 21-171 of PD-1 (SEQ ID NO:
327). In one embodiment, the antibodies may bind to an epitope
comprising one or more amino acids selected from the group
consisting of amino acid residues 1-146 of SEQ ID NOs: 321-324.
[0131] In certain embodiments, the antibodies of the present
invention may function by blocking or inhibiting the PD-L1-binding
activity associated with PD-1 by binding to any other region or
fragment of the full length protein, the amino acid sequence of
which is shown in SEQ ID NO: 327. In certain embodiments, the
antibodies may attenuate or modulate the interaction between PD-1
and PD-L1.
[0132] In certain embodiments, the antibodies of the present
invention may be bi-specific antibodies. The bi-specific antibodies
of the invention may bind one epitope in one domain and may also
bind a second epitope in a different domain of PD-1. In certain
embodiments, the bi-specific antibodies of the invention may bind
two different epitopes in the same domain. In one embodiment, the
multi-specific antigen-binding molecule comprises a first binding
specificity wherein the first binding specificity comprises the
extracellular domain or fragment thereof of PD-L1; and a second
binding specificity to another epitope of PD-1.
[0133] In one embodiment, the invention provides an isolated fully
human monoclonal antibody or antigen-binding fragment thereof that
binds to PD-1, wherein the antibody or fragment thereof exhibits
one or more of the following characteristics: (i) comprises a HCVR
having an amino acid sequence selected from the group consisting of
SEQ ID NO: 2, 18, 34, 50, 66, 82, 98, 114, 130, 146, 162, 178, 194,
210, 218, 226, 234, 242, 250, 258, 266, 274, 282, 290, 298, 306,
and 314, or a substantially similar sequence thereof having at
least 90%, at least 95%, at least 98% or at least 99% sequence
identity; (ii) comprises a LCVR having an amino acid sequence
selected from the group consisting of SEQ ID NO: 10, 26, 42, 58,
74, 90, 106, 122, 138, 154, 170, 186, and 202, or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity; (iii) comprises a
HCDR3 domain having an amino acid sequence selected from the group
consisting of SEQ ID NO: 8, 24, 40, 56, 72, 88, 104, 120, 136, 152,
168, 184, 200, 216, 224, 232, 240, 248, 256, 264, 272, 280, 288,
296, 304, 312, and 320, or a substantially similar sequence thereof
having at least 90%, at least 95%, at least 98% or at least 99%
sequence identity; and a LCDR3 domain having an amino acid sequence
selected from the group consisting of SEQ ID NO: 16, 32, 48, 64,
80, 96, 112, 128, 144, 160, 176, 192, and 208, or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity; (iv) comprises a HCDR1
domain having an amino acid sequence selected from the group
consisting of SEQ ID NO: 4, 20, 36, 52, 68, 84, 100, 116, 132, 148,
164, 180, 196, 212, 220, 228, 236, 244, 252, 260, 268, 276, 284,
292, 300, 308, and 316, or a substantially similar sequence thereof
having at least 90%, at least 95%, at least 98% or at least 99%
sequence identity; a HCDR2 domain having an amino acid sequence
selected from the group consisting of SEQ ID NO: 6, 22, 38, 54, 70,
86, 102, 118, 134, 150, 166, 182, 198, 214, 222, 230, 238, 246,
254, 262, 270, 278, 286, 294, 302, 310, and 318, or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity; a LCDR1 domain having
an amino acid sequence selected from the group consisting of SEQ ID
NO: 12, 28, 44, 60, 76, 92, 108, 124, 140, 156, 172, 188, and 204,
or a substantially similar sequence thereof having at least 90%, at
least 95%, at least 98% or at least 99% sequence identity; and a
LCDR2 domain having an amino acid sequence selected from the group
consisting of SEQ ID NO: 14, 30, 46, 62, 78, 94, 110, 126, 142,
158, 174, 190, and 206, or a substantially similar sequence thereof
having at least 90%, at least 95%, at least 98% or at least 99%
sequence identity; (v) is a multi-specific antigen-binding molecule
comprising a first binding specificity to PD-1 and a second binding
specificity to an antigen selected from the group consisting of
PD-1, a tumor specific antigen, an autoimmune tissue specific
antigen, a virally infected cell antigen, a different T-cell
co-inhibitor, T-cell receptor, and a Fc receptor; (vi) binds to
human PD-1 with a K.sub.D of about 28 pM to about 1.5 pM; (vii)
binds to cynomolgus PD-1 with a K.sub.D of about 3 nM to about 7.5
pM; (viii) blocks or enhances the binding of PD-1 to PD-L1 with an
IC50 about 3.3 nM; (ix) blocks PD-1-induced T-cell down regulation
and/or rescues T-cell signaling in a T-cell/APC luciferase reporter
assay; (x) stimulates T-cell proliferation and activity in a mixed
lymphocyte reaction (MLR) assay; (xi) induces IL-2 and/or
IFN.gamma. production in a MLR assay; and (xii) suppresses tumor
growth and increases survival in subjects with cancer.
[0134] In one embodiment, the invention provides an isolated fully
human monoclonal antibody or antigen-binding fragment thereof that
blocks PD-1 binding to PD-L1, wherein the antibody or fragment
thereof exhibits one or more of the following characteristics: (i)
comprises a HCVR having an amino acid sequence selected from the
group consisting of SEQ ID NO: 130, 162, 234 and 314, or a
substantially similar sequence thereof having at least 90%, at
least 95%, at least 98% or at least 99% sequence identity; (ii)
comprises a LCVR having an amino acid sequence selected from the
group consisting of SEQ ID NO: 138, 170, 186, and 202, or a
substantially similar sequence thereof having at least 90%, at
least 95%, at least 98% or at least 99% sequence identity; (iii)
comprises a HCDR3 domain having an amino acid sequence selected
from the group consisting of SEQ ID NO: 136, 168, 240, and 320, or
a substantially similar sequence thereof having at least 90%, at
least 95%, at least 98% or at least 99% sequence identity; and a
LCDR3 domain having an amino acid sequence selected from the group
consisting of SEQ ID NO: 144, 176, 192, and 208, or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity; (iv) comprises a HCDR1
domain having an amino acid sequence selected from the group
consisting of SEQ ID NO: 132, 164, 236, and 316, or a substantially
similar sequence thereof having at least 90%, at least 95%, at
least 98% or at least 99% sequence identity; a HCDR2 domain having
an amino acid sequence selected from the group consisting of SEQ ID
NO: 134, 166, 238, and 318, or a substantially similar sequence
thereof having at least 90%, at least 95%, at least 98% or at least
99% sequence identity; a LCDR1 domain having an amino acid sequence
selected from the group consisting of SEQ ID NO: 140, 172, 188, and
204, or a substantially similar sequence thereof having at least
90%, at least 95%, at least 98% or at least 99% sequence identity;
and a LCDR2 domain having an amino acid sequence selected from the
group consisting of SEQ ID NO: 142, 174, 190, and 206, or a
substantially similar sequence thereof having at least 90%, at
least 95%, at least 98% or at least 99% sequence identity; (v) is a
multi-specific antigen-binding molecule comprising a first binding
specificity to PD-1 and a second binding specificity to an antigen
selected from the group consisting of a different epitope of PD-1,
a tumor specific antigen, an autoimmune tissue specific antigen, a
virally infected cell antigen, a different T-cell co-inhibitor,
T-cell receptor, and a Fc receptor; (vi) binds to human PD-1 with a
K.sub.D.ltoreq.10.sup.-9M; (vii) binds to cynomolgus PD-1 with a
K.sub.D.ltoreq.10.sup.-8M; (viii) blocks the binding of PD-1 to
PD-L1 with an IC50.ltoreq.10.sup.-10M; (ix) blocks PD-1-induced
T-cell down regulation and/or rescues T-cell signaling in a
T-cell/APC luciferase reporter assay; (x) stimulates T-cell
proliferation and activity in a mixed lymphocyte reaction (MLR)
assay; (xi) induces IL-2 and/or IFN.gamma. production in a MLR
assay; and (xii) suppresses tumor growth and increases survival in
subjects with cancer.
[0135] The antibodies of the present invention may possess one or
more of the aforementioned biological characteristics, or any
combinations thereof. Other biological characteristics of the
antibodies of the present invention will be evident to a person of
ordinary skill in the art from a review of the present disclosure
including the working Examples herein.
Species Selectivity and Species Cross-Reactivity
[0136] According to certain embodiments of the invention, the
anti-PD-1 antibodies bind to human PD-1 but not to PD-1 from other
species. Alternatively, the anti-PD-1 antibodies of the invention,
in certain embodiments, bind to human PD-1 and to PD-1 from one or
more non-human species. For example, the anti-PD-1 antibodies of
the invention may bind to human PD-1 and may bind or not bind, as
the case may be, to one or more of mouse, rat, guinea pig, hamster,
gerbil, pig, cat, dog, rabbit, goat, sheep, cow, horse, camel,
cynomolgus, marmoset, rhesus or chimpanzee PD-1. In certain
embodiments, the anti-PD-1 antibodies of the invention may bind to
human and cynomolgus PD-1 with the same affinities or with
different affinities, but do not bind to rat and mouse PD-1.
Epitope Mapping and Related Technologies
[0137] The present invention includes anti-PD-1 antibodies which
interact with one or more amino acids found within one or more
domains of the PD-1 molecule including, e.g., extracellular
(IgV-like) domain, a transmembrane domain, and an intracellular
domain containing the immunoreceptor tyrosine-based inhibition
motif (ITIM) and immunoreceptor tyrosine-based switch motif (ITSM).
The epitope to which the antibodies bind may consist of a single
contiguous sequence of 3 or more (e.g., 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more) amino acids located
within any of the aforementioned domains of the PD-1 molecule (e.g.
a linear epitope in a domain). Alternatively, the epitope may
consist of a plurality of non-contiguous amino acids (or amino acid
sequences) located within either or both of the aforementioned
domains of the PD-1 molecule (e.g. a conformational epitope).
[0138] Various techniques known to persons of ordinary skill in the
art can be used to determine whether an antibody "interacts with
one or more amino acids" within a polypeptide or protein. Exemplary
techniques include, for example, routine cross-blocking assays,
such as that described in Antibodies, Harlow and Lane (Cold Spring
Harbor Press, Cold Spring Harbor, N.Y.). Other methods include
alanine scanning mutational analysis, peptide blot analysis
(Reineke (2004) Methods Mol. Biol. 248: 443-63), peptide cleavage
analysis crystallographic studies and NMR analysis. In addition,
methods such as epitope excision, epitope extraction and chemical
modification of antigens can be employed (Tomer (2000) Prot. Sci.
9: 487-496). Another method that can be used to identify the amino
acids within a polypeptide with which an antibody interacts is
hydrogen/deuterium exchange detected by mass spectrometry. In
general terms, the hydrogen/deuterium exchange method involves
deuterium-labeling the protein of interest, followed by binding the
antibody to the deuterium-labeled protein. Next, the
protein/antibody complex is transferred to water and exchangeable
protons within amino acids that are protected by the antibody
complex undergo deuterium-to-hydrogen back-exchange at a slower
rate than exchangeable protons within amino acids that are not part
of the interface. As a result, amino acids that form part of the
protein/antibody interface may retain deuterium and therefore
exhibit relatively higher mass compared to amino acids not included
in the interface. After dissociation of the antibody, the target
protein is subjected to protease cleavage and mass spectrometry
analysis, thereby revealing the deuterium-labeled residues which
correspond to the specific amino acids with which the antibody
interacts. See, e.g., Ehring (1999) Analytical Biochemistry 267:
252-259; Engen and Smith (2001) Anal. Chem. 73: 256A-265A.
[0139] The term "epitope" refers to a site on an antigen to which B
and/or T cells respond. B-cell epitopes can be formed both from
contiguous amino acids or noncontiguous amino acids juxtaposed by
tertiary folding of a protein. Epitopes formed from contiguous
amino acids are typically retained on exposure to denaturing
solvents, whereas epitopes formed by tertiary folding are typically
lost on treatment with denaturing solvents. An epitope typically
includes at least 3, and more usually, at least 5 or 8-10 amino
acids in a unique spatial conformation.
[0140] Modification-Assisted Profiling (MAP), also known as Antigen
Structure-based Antibody Profiling (ASAP) is a method that
categorizes large numbers of monoclonal antibodies (mAbs) directed
against the same antigen according to the similarities of the
binding profile of each antibody to chemically or enzymatically
modified antigen surfaces (see US 2004/0101920, herein specifically
incorporated by reference in its entirety). Each category may
reflect a unique epitope either distinctly different from or
partially overlapping with epitope represented by another category.
This technology allows rapid filtering of genetically identical
antibodies, such that characterization can be focused on
genetically distinct antibodies. When applied to hybridoma
screening, MAP may facilitate identification of rare hybridoma
clones that produce mAbs having the desired characteristics. MAP
may be used to sort the antibodies of the invention into groups of
antibodies binding different epitopes.
[0141] In certain embodiments, the anti-PD-1 antibodies or
antigen-binding fragments thereof bind an epitope within any one or
more of the regions exemplified in PD-1, either in natural form, as
exemplified in SEQ ID NO: 327, or recombinantly produced, as
exemplified in SEQ ID NOS: 321-324, or to a fragment thereof. In
some embodiments, the antibodies of the invention bind to an
extracellular region comprising one or more amino acids selected
from the group consisting of amino acid residues 21-171 of PD-1. In
some embodiments, the antibodies of the invention bind to an
extracellular region comprising one or more amino acids selected
from the group consisting of amino acid residues 1-146 of
cynomolgus PD-1, as exemplified by SEQ ID NO: 322.
[0142] In certain embodiments, the antibodies of the invention, as
shown in Table 1, interact with at least one amino acid sequence
selected from the group consisting of amino acid residues ranging
from about position 21 to about position 136 of SEQ ID NO: 327; or
amino acid residues ranging from about position 136 to about
position 171 of SEQ ID NO: 327. These regions are partially
exemplified in SEQ ID NOs: 321-324.
[0143] The present invention includes anti-PD-1 antibodies that
bind to the same epitope, or a portion of the epitope, as any of
the specific exemplary antibodies described herein in Table 1, or
an antibody having the CDR sequences of any of the exemplary
antibodies described in Table 1. Likewise, the present invention
also includes anti-PD-1 antibodies that compete for binding to PD-1
or a PD-1 fragment with any of the specific exemplary antibodies
described herein in Table 1, or an antibody having the CDR
sequences of any of the exemplary antibodies described in Table 1.
For example, the present invention includes anti-PD-1 antibodies
that cross-compete for binding to PD-1 with one or more antibodies
as defined in Example 6 herein (e.g., H2aM7788N, H4xH8992P,
H4xH8999P, H1M7799N, H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P,
H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P,
H4xH9045P and H2aM7795N).
[0144] One can easily determine whether an antibody binds to the
same epitope as, or competes for binding with, a reference
anti-PD-1 antibody by using routine methods known in the art. For
example, to determine if a test antibody binds to the same epitope
as a reference anti-PD-1 antibody of the invention, the reference
antibody is allowed to bind to a PD-1 protein or peptide under
saturating conditions. Next, the ability of a test antibody to bind
to the PD-1 molecule is assessed. If the test antibody is able to
bind to PD-1 following saturation binding with the reference
anti-PD-1 antibody, it can be concluded that the test antibody
binds to a different epitope than the reference anti-PD-1 antibody.
On the other hand, if the test antibody is not able to bind to the
PD-1 protein following saturation binding with the reference
anti-PD-1 antibody, then the test antibody may bind to the same
epitope as the epitope bound by the reference anti-PD-1 antibody of
the invention.
[0145] To determine if an antibody competes for binding with a
reference anti-PD-1 antibody, the above-described binding
methodology is performed in two orientations: In a first
orientation, the reference antibody is allowed to bind to a PD-1
protein under saturating conditions followed by assessment of
binding of the test antibody to the PD-1 molecule. In a second
orientation, the test antibody is allowed to bind to a PD-1
molecule under saturating conditions followed by assessment of
binding of the reference antibody to the PD-1 molecule. If, in both
orientations, only the first (saturating) antibody is capable of
binding to the PD-1 molecule, then it is concluded that the test
antibody and the reference antibody compete for binding to PD-1. As
will be appreciated by a person of ordinary skill in the art, an
antibody that competes for binding with a reference antibody may
not necessarily bind to the identical epitope as the reference
antibody, but may sterically block binding of the reference
antibody by binding an overlapping or adjacent epitope.
[0146] Two antibodies bind to the same or overlapping epitope if
each competitively inhibits (blocks) binding of the other to the
antigen. That is, a 1-, 5-, 10-, 20- or 100-fold excess of one
antibody inhibits binding of the other by at least 50% but
preferably 75%, 90% or even 99% as measured in a competitive
binding assay (see, e.g., Junghans et al., Cancer Res. 1990
50:1495-1502). Alternatively, two antibodies have the same epitope
if essentially all amino acid mutations in the antigen that reduce
or eliminate binding of one antibody reduce or eliminate binding of
the other. Two antibodies have overlapping epitopes if some amino
acid mutations that reduce or eliminate binding of one antibody
reduce or eliminate binding of the other.
[0147] Additional routine experimentation (e.g., peptide mutation
and binding analyses) can then be carried out to confirm whether
the observed lack of binding of the test antibody is in fact due to
binding to the same epitope as the reference antibody or if steric
blocking (or another phenomenon) is responsible for the lack of
observed binding. Experiments of this sort can be performed using
ELISA, RIA, surface plasmon resonance, flow cytometry or any other
quantitative or qualitative antibody-binding assay available in the
art.
Immunoconjugates
[0148] The invention encompasses a human anti-PD-1 monoclonal
antibody conjugated to a therapeutic moiety ("immunoconjugate"),
such as a cytotoxin or a chemotherapeutic agent to treat cancer. As
used herein, the term "immunoconjugate" refers to an antibody which
is chemically or biologically linked to a cytotoxin, a radioactive
agent, a cytokine, an interferon, a target or reporter moiety, an
enzyme, a toxin, a peptide or protein or a therapeutic agent. The
antibody may be linked to the cytotoxin, radioactive agent,
cytokine, interferon, target or reporter moiety, enzyme, toxin,
peptide or therapeutic agent at any location along the molecule so
long as it is able to bind its target. Examples of immunoconjugates
include antibody drug conjugates and antibody-toxin fusion
proteins. In one embodiment, the agent may be a second different
antibody to PD-1. In certain embodiments, the antibody may be
conjugated to an agent specific for a tumor cell or a virally
infected cell. The type of therapeutic moiety that may be
conjugated to the anti-PD-1 antibody and will take into account the
condition to be treated and the desired therapeutic effect to be
achieved. Examples of suitable agents for forming immunoconjugates
are known in the art; see for example, WO 05/103081.
Multi-Specific Antibodies
[0149] The antibodies of the present invention may be
mono-specific, bi-specific, or multi-specific. Multi-specific
antibodies may be specific for different epitopes of one target
polypeptide or may contain antigen-binding domains specific for
more than one target polypeptide. See, e.g., Tutt et al., 1991, J.
Immunol. 147:60-69; Kufer et al., 2004, Trends Biotechnol.
22:238-244.
[0150] In one aspect, the present invention includes multi-specific
antigen-binding molecules or antigen-binding fragments thereof
wherein one specificity of an immunoglobulin is specific for the
extracellular domain of PD-1, or a fragment thereof, and the other
specificity of the immunoglobulin is specific for binding outside
the extracellular domain of PD-1, or a second therapeutic target,
or is conjugated to a therapeutic moiety. In certain embodiments,
the first antigen-binding specificity may comprise PD-L1 or PD-L2,
or a fragment thereof. In certain embodiments of the invention, one
specificity of an immunoglobulin is specific for an epitope
comprising amino acid residues 21-171 of PD-1 (SEQ ID NO: 327) or a
fragment thereof, and the other specificity of the immunoglobulin
is specific for a second target antigen. The second target antigen
may be on the same cell as PD-1 or on a different cell. In one
embodiment, the second target cell is on an immune cell other than
a T-cell such as a B-cell, antigen-presenting cell, monocyte,
macrophage, or dendritic cell. In some embodiments, the second
target antigen may be present on a tumor cell or an autoimmune
tissue cell or on a virally infected cell.
[0151] In another aspect, the invention provides multi-specific
antigen-binding molecules or antigen-binding fragments thereof
comprising a first antigen-binding specificity that binds to PD-1
and a second antigen-binding specificity that binds to a T-cell
receptor, a B-cell receptor or a Fc receptor. In a related aspect,
the invention provides multi-specific antigen-binding molecules or
antigen-binding fragments thereof comprising a first
antigen-binding specificity that binds to PD-1 and a second
antigen-binding specificity that binds to a different T-cell
co-inhibitor such as LAG-3, CTLA-4, BTLA, CD-28, 2B4, LY108, TIGIT,
TIM3, LAIR1, ICOS and CD160.
[0152] In another aspect, the invention provides multi-specific
antigen-binding molecules or antigen-binding fragments thereof
comprising a first antigen-binding specificity that binds to PD-1
and a second antigen-binding specificity that binds to an
autoimmune tissue-specific antigen. In certain embodiments, the
antibodies may be activating or agonist antibodies.
[0153] Any of the multi-specific antigen-binding molecules of the
invention, or variants thereof, may be constructed using standard
molecular biological techniques (e.g., recombinant DNA and protein
expression technology), as will be known to a person of ordinary
skill in the art.
[0154] In some embodiments, PD-1-specific antibodies are generated
in a bi-specific format (a "bi-specific") in which variable regions
binding to distinct domains of PD-1 are linked together to confer
dual-domain specificity within a single binding molecule.
Appropriately designed bi-specifics may enhance overall PD-1
inhibitory efficacy through increasing both specificity and binding
avidity. Variable regions with specificity for individual domains,
(e.g., segments of the N-terminal domain), or that can bind to
different regions within one domain, are paired on a structural
scaffold that allows each region to bind simultaneously to the
separate epitopes, or to different regions within one domain. In
one example for a bi-specific, heavy chain variable regions
(V.sub.H) from a binder with specificity for one domain are
recombined with light chain variable regions (V.sub.L) from a
series of binders with specificity for a second domain to identify
non-cognate V.sub.L partners that can be paired with an original
V.sub.H without disrupting the original specificity for that
V.sub.H. In this way, a single V.sub.L segment (e.g., V.sub.L1) can
be combined with two different V.sub.H domains (e.g., V.sub.H1 and
V.sub.H2) to generate a bi-specific comprised of two binding "arms"
(V.sub.H1-V.sub.L1 and V.sub.H2-V.sub.L1). Use of a single V.sub.L
segment reduces the complexity of the system and thereby simplifies
and increases efficiency in cloning, expression, and purification
processes used to generate the bi-specific (See, for example, U.S.
Ser. No. 13/022,759 and US2010/0331527).
[0155] Alternatively, antibodies that bind more than one domains
and a second target, such as, but not limited to, for example, a
second different anti-PD-1 antibody, may be prepared in a
bi-specific format using techniques described herein, or other
techniques known to those skilled in the art. Antibody variable
regions binding to distinct regions may be linked together with
variable regions that bind to relevant sites on, for example, the
extracellular domain of PD-1, to confer dual-antigen specificity
within a single binding molecule. Appropriately designed
bi-specifics of this nature serve a dual function. Variable regions
with specificity for the extracellular domain are combined with a
variable region with specificity for outside the extracellular
domain and are paired on a structural scaffold that allows each
variable region to bind to the separate antigens.
[0156] An exemplary bi-specific antibody format that can be used in
the context of the present invention involves the use of a first
immunoglobulin (Ig) C.sub.H3 domain and a second Ig C.sub.H3
domain, wherein the first and second Ig C.sub.H3 domains differ
from one another by at least one amino acid, and wherein at least
one amino acid difference reduces binding of the bi-specific
antibody to Protein A as compared to a bi-specific antibody lacking
the amino acid difference. In one embodiment, the first Ig C.sub.H3
domain binds Protein A and the second Ig C.sub.H3 domain contains a
mutation that reduces or abolishes Protein A binding such as an
H95R modification (by IMGT exon numbering; H435R by EU numbering).
The second C.sub.H3 may further comprise a Y96F modification (by
IMGT; Y436F by EU). Further modifications that may be found within
the second C.sub.H3 include: D16E, L18M, N44S, K52N, V57M, and V82I
(by IMGT; D356E, L358M, N384S, K392N, V397M, and V422I by EU) in
the case of IgG1 antibodies; N44S, K52N, and V82I (IMGT; N384S,
K392N, and V422I by EU) in the case of IgG2 antibodies; and Q15R,
N44S, K52N, V57M, R69K, E79Q, and V82I (by IMGT; Q355R, N384S,
K392N, V397M, R409K, E419Q, and V422I by EU) in the case of IgG4
antibodies. Variations on the bi-specific antibody format described
above are contemplated within the scope of the present
invention.
[0157] Other exemplary bispecific formats that can be used in the
context of the present invention include, without limitation, e.g.,
scFv-based or diabody bispecific formats, IgG-scFv fusions, dual
variable domain (DVD)-Ig, Quadroma, knobs-into-holes, common light
chain (e.g., common light chain with knobs-into-holes, etc.),
CrossMab, CrossFab, (SEED)body, leucine zipper, Duobody, IgG1/IgG2,
dual acting Fab (DAF)-IgG, and Mabe bispecific formats (see, e.g.,
Klein et al. 2012, mAbs 4:6, 1-11, and references cited therein,
for a review of the foregoing formats). Bispecific antibodies can
also be constructed using peptide/nucleic acid conjugation, e.g.,
wherein unnatural amino acids with orthogonal chemical reactivity
are used to generate site-specific antibody-oligonucleotide
conjugates which then self-assemble into multimeric complexes with
defined composition, valency and geometry. (See, e.g., Kazane et
al., J. Am. Chem. Soc. [Epub: Dec. 4, 2012]).
Therapeutic Administration and Formulations
[0158] The invention provides therapeutic compositions comprising
the anti-PD-1 antibodies or antigen-binding fragments thereof of
the present invention. Therapeutic compositions in accordance with
the invention will be administered with suitable carriers,
excipients, and other agents that are incorporated into
formulations to provide improved transfer, delivery, tolerance, and
the like. A multitude of appropriate formulations can be found in
the formulary known to all pharmaceutical chemists: Remington's
Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa. These
formulations include, for example, powders, pastes, ointments,
jellies, waxes, oils, lipids, lipid (cationic or anionic)
containing vesicles (such as LIPOFECTIN.TM.), DNA conjugates,
anhydrous absorption pastes, oil-in-water and water-in-oil
emulsions, emulsions carbowax (polyethylene glycols of various
molecular weights), semi-solid gels, and semi-solid mixtures
containing carbowax. See also Powell et al. "Compendium of
excipients for parenteral formulations" PDA (1998) J Pharm Sci
Technol 52:238-311.
[0159] The dose of antibody may vary depending upon the age and the
size of a subject to be administered, target disease, conditions,
route of administration, and the like. When an antibody of the
present invention is used for treating a disease or disorder in an
adult patient, or for preventing such a disease, it is advantageous
to administer the antibody of the present invention normally at a
single dose of about 0.1 to about 60 mg/kg body weight, more
preferably about 5 to about 60, about 10 to about 50, or about 20
to about 50 mg/kg body weight. Depending on the severity of the
condition, the frequency and the duration of the treatment can be
adjusted. In certain embodiments, the antibody or antigen-binding
fragment thereof of the invention can be administered as an initial
dose of at least about 0.1 mg to about 800 mg, about 1 to about 500
mg, about 5 to about 300 mg, or about 10 to about 200 mg, to about
100 mg, or to about 50 mg. In certain embodiments, the initial dose
may be followed by administration of a second or a plurality of
subsequent doses of the antibody or antigen-binding fragment
thereof in an amount that can be approximately the same or less
than that of the initial dose, wherein the subsequent doses are
separated by at least 1 day to 3 days; at least one week, at least
2 weeks; at least 3 weeks; at least 4 weeks; at least 5 weeks; at
least 6 weeks; at least 7 weeks; at least 8 weeks; at least 9
weeks; at least 10 weeks; at least 12 weeks; or at least 14
weeks.
[0160] Various delivery systems are known and can be used to
administer the pharmaceutical composition of the invention, e.g.,
encapsulation in liposomes, microparticles, microcapsules,
recombinant cells capable of expressing the mutant viruses,
receptor mediated endocytosis (see, e.g., Wu et al. (1987) J. Biol.
Chem. 262:4429-4432). Methods of introduction include, but are not
limited to, intradermal, transdermal, intramuscular,
intraperitoneal, intravenous, subcutaneous, intranasal, epidural
and oral routes. The composition may be administered by any
convenient route, for example by infusion or bolus injection, by
absorption through epithelial or mucocutaneous linings (e.g., oral
mucosa, rectal and intestinal mucosa, etc.) and may be administered
together with other biologically active agents. Administration can
be systemic or local. The pharmaceutical composition can be also
delivered in a vesicle, in particular a liposome (see, for example,
Langer (1990) Science 249:1527-1533).
[0161] The use of nanoparticles to deliver the antibodies of the
present invention is also contemplated herein. Antibody-conjugated
nanoparticles may be used both for therapeutic and diagnostic
applications. Antibody-conjugated nanoparticles and methods of
preparation and use are described in detail by Arruebo, M., et al.
2009 ("Antibody-conjugated nanoparticles for biomedical
applications" in J. Nanomat. Volume 2009, Article ID 439389, 24
pages, doi: 10.1155/2009/439389), incorporated herein by reference.
Nanoparticles may be developed and conjugated to antibodies
contained in pharmaceutical compositions to target tumor cells or
autoimmune tissue cells or virally infected cells. Nanoparticles
for drug delivery have also been described in, for example, U.S.
Pat. No. 8,257,740, or U.S. Pat. No. 8,246,995, each incorporated
herein in its entirety.
[0162] In certain situations, the pharmaceutical composition can be
delivered in a controlled release system. In one embodiment, a pump
may be used. In another embodiment, polymeric materials can be
used. In yet another embodiment, a controlled release system can be
placed in proximity of the composition's target, thus requiring
only a fraction of the systemic dose.
[0163] The injectable preparations may include dosage forms for
intravenous, subcutaneous, intracutaneous, intracranial,
intraperitoneal and intramuscular injections, drip infusions, etc.
These injectable preparations may be prepared by methods publicly
known. For example, the injectable preparations may be prepared,
e.g., by dissolving, suspending or emulsifying the antibody or its
salt described above in a sterile aqueous medium or an oily medium
conventionally used for injections. As the aqueous medium for
injections, there are, for example, physiological saline, an
isotonic solution containing glucose and other auxiliary agents,
etc., which may be used in combination with an appropriate
solubilizing agent such as an alcohol (e.g., ethanol), a
polyalcohol (e.g., propylene glycol, polyethylene glycol), a
nonionic surfactant [e.g., polysorbate 80, HCO-50 (polyoxyethylene
(50 mol) adduct of hydrogenated castor oil)], etc. As the oily
medium, there are employed, e.g., sesame oil, soybean oil, etc.,
which may be used in combination with a solubilizing agent such as
benzyl benzoate, benzyl alcohol, etc. The injection thus prepared
is preferably filled in an appropriate ampoule.
[0164] A pharmaceutical composition of the present invention can be
delivered subcutaneously or intravenously with a standard needle
and syringe. In addition, with respect to subcutaneous delivery, a
pen delivery device readily has applications in delivering a
pharmaceutical composition of the present invention. Such a pen
delivery device can be reusable or disposable. A reusable pen
delivery device generally utilizes a replaceable cartridge that
contains a pharmaceutical composition. Once all of the
pharmaceutical composition within the cartridge has been
administered and the cartridge is empty, the empty cartridge can
readily be discarded and replaced with a new cartridge that
contains the pharmaceutical composition. The pen delivery device
can then be reused. In a disposable pen delivery device, there is
no replaceable cartridge. Rather, the disposable pen delivery
device comes prefilled with the pharmaceutical composition held in
a reservoir within the device. Once the reservoir is emptied of the
pharmaceutical composition, the entire device is discarded.
[0165] Numerous reusable pen and autoinjector delivery devices have
applications in the subcutaneous delivery of a pharmaceutical
composition of the present invention. Examples include, but
certainly are not limited to AUTOPEN.TM. (Owen Mumford, Inc.,
Woodstock, UK), DISETRONIC.TM. pen (Disetronic Medical Systems,
Burghdorf, Switzerland), HUMALOG MIX 75/25.TM. pen, HUMALOG.TM.
pen, HUMALIN 70/30.TM. pen (Eli Lilly and Co., Indianapolis, Ind.),
NOVOPEN.TM. I, II and III (Novo Nordisk, Copenhagen, Denmark),
NOVOPEN JUNIOR.TM. (Novo Nordisk, Copenhagen, Denmark), BD.TM. pen
(Becton Dickinson, Franklin Lakes, N.J.), OPTIPEN.TM., OPTIPEN
PRO.TM., OPTIPEN STARLET.TM., and OPTICLIK.TM. (Sanofi-Aventis,
Frankfurt, Germany), to name only a few. Examples of disposable pen
delivery devices having applications in subcutaneous delivery of a
pharmaceutical composition of the present invention include, but
certainly are not limited to the SOLOSTAR.TM. pen (Sanofi-Aventis),
the FLEXPEN.TM. (Novo Nordisk), and the KWIKPEN.TM. (Eli Lilly),
the SURECLICK.TM. Autoinjector (Amgen, Thousand Oaks, Calif.), the
PENLET.TM. (Haselmeier, Stuttgart, Germany), the EPIPEN (Dey, L.P.)
and the HUMIRA.TM. Pen (Abbott Labs, Abbott Park, Ill.), to name
only a few.
[0166] Advantageously, the pharmaceutical compositions for oral or
parenteral use described above are prepared into dosage forms in a
unit dose suited to fit a dose of the active ingredients. Such
dosage forms in a unit dose include, for example, tablets, pills,
capsules, injections (ampoules), suppositories, etc. The amount of
the antibody contained is generally about 5 to about 500 mg per
dosage form in a unit dose; especially in the form of injection, it
is preferred that the antibody is contained in about 5 to about 100
mg and in about 10 to about 250 mg for the other dosage forms.
Therapeutic Uses of the Antibodies
[0167] The antibodies of the invention are useful, inter alia, for
the treatment, prevention and/or amelioration of any disease or
disorder associated with or mediated by PD-1 expression, signaling,
or activity, or treatable by blocking the interaction between PD-1
and a PD-1 ligand (e.g., PD-L1, or PD-L2) or otherwise inhibiting
PD-1 activity and/or signaling. For example, the present invention
provides methods for treating cancer (tumor growth inhibition),
chronic viral infections and/or autoimmune disease by administering
an anti-PD-1 antibody (or pharmaceutical composition comprising an
anti-PD-1 antibody) as described herein to a patient in need of
such treatment. The antibodies of the present invention are useful
for the treatment, prevention, and/or amelioration of disease or
disorder or condition such as cancer, autoimmune disease or a viral
infection and/or for ameliorating at least one symptom associated
with such disease, disorder or condition. In the context of the
methods of treatment described herein, the anti-PD-1 antibody may
be administered as a monotherapy (i.e., as the only therapeutic
agent) or in combination with one or more additional therapeutic
agents (examples of which are described elsewhere herein).
[0168] In some embodiments of the invention, the antibodies
described herein are useful for treating subjects suffering from
primary or recurrent cancer, including, but not limited to, renal
cell carcinoma, colorectal cancer, non-small-cell lung cancer,
brain cancer (e.g., glioblastoma multiforme), squamous cell
carcinoma of head and neck, gastric cancer, prostate cancer,
ovarian cancer, kidney cancer, breast cancer, multiple myeloma, and
melanoma.
[0169] The antibodies may be used to treat early stage or
late-stage symptoms of cancer. In one embodiment, an antibody or
fragment thereof of the invention may be used to treat metastatic
cancer. The antibodies are useful in reducing or inhibiting or
shrinking tumor growth of both solid tumors and blood cancers. In
certain embodiments, treatment with an antibody or antigen-binding
fragment thereof of the invention leads to more than 50%
regression, more than 60% regression, more than 70% regression,
more than 80% regression or more than 90% regression of a tumor in
a subject. In certain embodiments, the antibodies may be used to
prevent relapse of a tumor. In certain embodiments, the antibodies
are useful in extending overall survival in a subject with cancer.
In some embodiments, the antibodies are useful in reducing toxicity
due to chemotherapy or radiotherapy while maintaining long-term
survival in a patient suffering from cancer.
[0170] In certain embodiments, the antibodies of the invention are
useful to treat subjects suffering from a chronic viral infection.
In some embodiments, the antibodies of the invention are useful in
decreasing viral titers in the host and/or rescuing exhausted
T-cells. In certain embodiments, an antibody or fragment thereof of
the invention may be used to treat chronic viral infection by
lymphocytic choriomeningitis virus (LCMV). In some embodiments, an
antibody or antigen-binding fragment thereof the invention may be
administered at a therapeutic dose to a patient with an infection
by human immunodeficiency virus (HIV) or human papilloma virus
(HPV) or hepatitis B/C virus (HBV/HCV). In a related embodiment, an
antibody or antigen-binding fragment thereof of the invention may
be used to treat an infection by simian immunodeficiency virus
(SIV) in a simian subject such as cynomolgus.
[0171] In certain embodiments, a blocking antibody of the present
invention may be administered in a therapeutically effective amount
to a subject suffering from a cancer or a viral infection.
[0172] In certain embodiments, the antibodies of the invention are
useful for treating an autoimmune disease, including but not
limited to, alopecia areata, autoimmune hepatitis, celiac disease,
Graves' disease, Guillain-Barre syndrome, Hashimoto's disease,
hemolytic anemia, inflammatory bowel disease, inflammatory
myopathies, multiple sclerosis, primary biliary cirrhosis,
psoriasis, rheumatoid arthritis, scleroderma, Sjogren's syndrome,
systemic lupus erthyematosus, vitiligo, autoimmune pancreatitis,
autoimmune urticaria, autoimmune thrombocytopenic purpura, Crohn's
disease, diabetes type I, eosinophilic fasciitis, eosinophilic
enterogastritis, Goodpasture's syndrome, myasthenia gravis,
psoriatic arthritis, rheumatic fever, ulcerative colitis,
vasculitis and Wegener's granulomatosis. In certain embodiments, an
activating antibody of the invention may be used to treat a subject
suffering from autoimmune disease.
[0173] One or more antibodies of the present invention may be
administered to relieve or prevent or decrease the severity of one
or more of the symptoms or conditions of the disease or
disorder.
[0174] It is also contemplated herein to use one or more antibodies
of the present invention prophylactically to patients at risk for
developing a disease or disorder such as cancer, autoimmune disease
and chronic viral infection.
[0175] In a further embodiment of the invention the present
antibodies are used for the preparation of a pharmaceutical
composition for treating patients suffering from cancer, autoimmune
disease or viral infection. In another embodiment of the invention,
the present antibodies are used as adjunct therapy with any other
agent or any other therapy known to those skilled in the art useful
for treating cancer, autoimmune disease or viral infection.
Combination Therapies and Formulations
[0176] Combination therapies may include an anti-PD-1 antibody of
the invention and any additional therapeutic agent that may be
advantageously combined with an antibody of the invention, or with
a biologically active fragment of an antibody of the invention.
[0177] The antibodies of the present invention may be combined
synergistically with one or more anti-cancer drugs or therapy used
to treat cancer, including, for example, renal cell carcinoma,
colorectal cancer, glioblastoma multiforme, squamous cell carcinoma
of head and neck, non-small-cell lung cancer, colon cancer, ovarian
cancer, adenocarcinoma, prostate cancer, glioma, and melanoma. It
is contemplated herein to use anti-PD-1 antibodies of the invention
in combination with immunostimulatory and/or immunosupportive
therapies to inhibit tumor growth, and/or enhance survival of
cancer patients. The immunostimulatory therapies include direct
immunostimulatory therapies to augment immune cell activity by
either "releasing the brake" on suppressed immune cells or
"stepping on the gas" to activate an immune response. Examples
include targeting other checkpoint receptors, vaccination and
adjuvants. The immunosupportive modalities may increase
antigenicity of the tumor by promoting immunogenic cell death,
inflammation or have other indirect effects that promote an
anti-tumor immune response. Examples include radiation,
chemotherapy, anti-angiogenic agents, and surgery.
[0178] In various embodiments, one or more antibodies of the
present invention may be used in combination with an antibody to
PD-L1, a second antibody to PD-1 (e.g., nivolumab), a LAG-3
inhibitor, a CTLA-4 inhibitor (e.g., ipilimumab), a TIM3 inhibitor,
a BTLA inhibitor, a TIGIT inhibitor, a CD47 inhibitor, an
antagonist of another T-cell co-inhibitor or ligand (e.g., an
antibody to CD-28, 2B4, LY108, LAIR1, ICOS, CD160 or VISTA), an
indoleamine-2,3-dioxygenase (IDO) inhibitor, a vascular endothelial
growth factor (VEGF) antagonist [e.g., a "VEGF-Trap" such as
aflibercept or other VEGF-inhibiting fusion protein as set forth in
U.S. Pat. No. 7,087,411, or an anti-VEGF antibody or antigen
binding fragment thereof (e.g., bevacizumab, or ranibizumab) or a
small molecule kinase inhibitor of VEGF receptor (e.g., sunitinib,
sorafenib, or pazopanib)], an Ang2 inhibitor (e.g., nesvacumab), a
transforming growth factor beta (TGF.beta.) inhibitor, an epidermal
growth factor receptor (EGFR) inhibitor (e.g., erlotinib,
cetuximab), an agonist to a co-stimulatory receptor (e.g., an
agonist to glucocorticoid-induced TNFR-related protein), an
antibody to a tumor-specific antigen (e.g., CA9, CA125,
melanoma-associated antigen 3 (MAGE3), carcinoembryonic antigen
(CEA), vimentin, tumor-M2-PK, prostate-specific antigen (PSA),
mucin-1, MART-1, and CA19-9), a vaccine (e.g., Bacillus
Calmette-Guerin, a cancer vaccine), an adjuvant to increase antigen
presentation (e.g., granulocyte-macrophage colony-stimulating
factor), a bispecific antibody (e.g., CD3xCD20 bispecific antibody,
PSMAxCD3 bispecific antibody), a cytotoxin, a chemotherapeutic
agent (e.g., dacarbazine, temozolomide, cyclophosphamide,
docetaxel, doxorubicin, daunorubicin, cisplatin, carboplatin,
gemcitabine, methotrexate, mitoxantrone, oxaliplatin, paclitaxel,
and vincristine), cyclophosphamide, radiotherapy, an IL-6R
inhibitor (e.g., sarilumab), an IL-4R inhibitor (e.g., dupilumab),
an IL-10 inhibitor, a cytokine such as IL-2, IL-7, IL-21, and
IL-15, an antibody-drug conjugate (ADC) (e.g., anti-CD19-DM4 ADC,
and anti-DS6-DM4 ADC), an anti-inflammatory drug (e.g.,
corticosteroids, and non-steroidal anti-inflammatory drugs), a
dietary supplement such as anti-oxidants or any palliative care to
treat cancer. In certain embodiments, the anti-PD-1 antibodies of
the present invention may be used in combination with cancer
vaccines including dendritic cell vaccines, oncolytic viruses,
tumor cell vaccines, etc. to augment the anti-tumor response.
Examples of cancer vaccines that can be used in combination with
anti-PD-1 antibodies of the present invention include MAGE3 vaccine
for melanoma and bladder cancer, MUC1 vaccine for breast cancer,
EGFRv3 (e.g., Rindopepimut) for brain cancer (including
glioblastoma multiforme), or ALVAC-CEA (for CEA+ cancers).
[0179] In certain embodiments, the anti-PD-1 antibodies of the
invention may be administered in combination with radiation therapy
in methods to generate long-term durable anti-tumor responses
and/or enhance survival of patients with cancer. In some
embodiments, the anti-PD-1 antibodies of the invention may be
administered prior to, concomitantly or after administering
radiation therapy to a cancer patient. For example, radiation
therapy may be administered in one or more doses to tumor lesions
followed by administration of one or more doses of anti-PD-1
antibodies of the invention. In some embodiments, radiation therapy
may be administered locally to a tumor lesion to enhance the local
immunogenicity of a patient's tumor (adjuvinating radiation) and/or
to kill tumor cells (ablative radiation) followed by systemic
administration of an anti-PD-1 antibody of the invention. For
example, intracranial radiation may be administered to a patient
with brain cancer (e.g., glioblastoma multiforme) in combination
with systemic administration of an anti-PD-1 antibody of the
invention. In certain embodiments, the anti-PD-1 antibodies of the
invention may be administered in combination with radiation therapy
and a chemotherapeutic agent (e.g., temozolomide) or a VEGF
antagonist (e.g., aflibercept).
[0180] In certain embodiments, the anti-PD-1 antibodies of the
invention may be administered in combination with one or more
anti-viral drugs to treat chronic viral infection caused by LCMV,
HIV, HPV, HBV or HCV. Examples of anti-viral drugs include, but are
not limited to, zidovudine, lamivudine, abacavir, ribavirin,
lopinavir, efavirenz, cobicistat, tenofovir, rilpivirine and
corticosteroids. In some embodiments, the anti-PD-1 antibodies of
the invention may be administered in combination with a LAG3
inhibitor, a CTLA-4 inhibitor or any antagonist of another T-cell
co-inhibitor to treat chronic viral infection.
[0181] In certain embodiments, the anti-PD-1 antibodies of the
invention may be combined with an antibody to a Fc receptor on
immune cells for the treatment of an autoimmune disease. In one
embodiment, an antibody or fragment thereof of the invention is
administered in combination with an antibody or antigen-binding
protein targeted to an antigen specific to autoimmune tissue. In
certain embodiments, an antibody or antigen-binding fragment
thereof of the invention is administered in combination with an
antibody or antigen-binding protein targeted to a T-cell receptor
or a B-cell receptor, including but not limited to, Fc.alpha.
(e.g., CD89), Fc.gamma. (e.g., CD64, CD32, CD16a, and CD16b), CD19,
etc. The antibodies of fragments thereof of the invention may be
used in combination with any drug or therapy known in the art
(e.g., corticosteroids and other immunosuppressants) to treat an
autoimmune disease or disorder including, but not limited to
alopecia areata, autoimmune hepatitis, celiac disease, Graves'
disease, Guillain-Barre syndrome, Hashimoto's disease, hemolytic
anemia, inflammatory bowel disease, inflammatory myopathies,
multiple sclerosis, primary biliary cirrhosis, psoriasis,
rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic
lupus erthyematosus, vitiligo, autoimmune pancreatitis, autoimmune
urticaria, autoimmune thrombocytopenic purpura, Crohn's disease,
diabetes type I, eosinophilic fasciitis, eosinophilic
enterogastritis, Goodpasture's syndrome, myasthenia gravis,
psoriatic arthritis, rheumatic fever, ulcerative colitis,
vasculitis and Wegener's granulomatosis.
[0182] The additional therapeutically active agent(s)/component(s)
may be administered prior to, concurrent with, or after the
administration of the anti-PD-1 antibody of the present invention.
For purposes of the present disclosure, such administration
regimens are considered the administration of an anti-PD-1 antibody
"in combination with" a second therapeutically active
component.
[0183] The additional therapeutically active component(s) may be
administered to a subject prior to administration of an anti-PD-1
antibody of the present invention. For example, a first component
may be deemed to be administered "prior to" a second component if
the first component is administered 1 week before, 72 hours before,
60 hours before, 48 hours before, 36 hours before, 24 hours before,
12 hours before, 6 hours before, 5 hours before, 4 hours before, 3
hours before, 2 hours before, 1 hour before, 30 minutes before, 15
minutes before, 10 minutes before, 5 minutes before, or less than 1
minute before administration of the second component. In other
embodiments, the additional therapeutically active component(s) may
be administered to a subject after administration of an anti-PD-1
antibody of the present invention. For example, a first component
may be deemed to be administered "after" a second component if the
first component is administered 1 minute after, 5 minutes after, 10
minutes after, 15 minutes after, 30 minutes after, 1 hour after, 2
hours after, 3 hours after, 4 hours after, 5 hours after, 6 hours
after, 12 hours after, 24 hours after, 36 hours after, 48 hours
after, 60 hours after, 72 hours after administration of the second
component. In yet other embodiments, the additional therapeutically
active component(s) may be administered to a subject concurrent
with administration of an anti-PD-1 antibody of the present
invention. "Concurrent" administration, for purposes of the present
invention, includes, e.g., administration of an anti-PD-1 antibody
and an additional therapeutically active component to a subject in
a single dosage form (e.g., co-formulated), or in separate dosage
forms administered to the subject within about 30 minutes or less
of each other. If administered in separate dosage forms, each
dosage form may be administered via the same route (e.g., both the
anti-PD-1 antibody and the additional therapeutically active
component may be administered intravenously, subcutaneously, etc.);
alternatively, each dosage form may be administered via a different
route (e.g., the anti-PD-1 antibody may be administered
intravenously, and the additional therapeutically active component
may be administered subcutaneously). In any event, administering
the components in a single dosage from, in separate dosage forms by
the same route, or in separate dosage forms by different routes are
all considered "concurrent administration," for purposes of the
present disclosure. For purposes of the present disclosure,
administration of an anti-PD-1 antibody "prior to", "concurrent
with," or "after" (as those terms are defined herein above)
administration of an additional therapeutically active component is
considered administration of an anti-PD-1 antibody "in combination
with" an additional therapeutically active component).
[0184] The present invention includes pharmaceutical compositions
in which an anti-PD-1 antibody of the present invention is
co-formulated with one or more of the additional therapeutically
active component(s) as described elsewhere herein using a variety
of dosage combinations.
[0185] In exemplary embodiments in which an anti-PD-1 antibody of
the invention is administered in combination with a VEGF antagonist
(e.g., a VEGF trap such as aflibercept), including administration
of co-formulations comprising an anti-PD-1 antibody and a VEGF
antagonist, the individual components may be administered to a
subject and/or co-formulated using a variety of dosage
combinations. For example, the anti-PD-1 antibody may be
administered to a subject and/or contained in a co-formulation in
an amount selected from the group consisting of 0.01 mg, 0.02 mg,
0.03 mg, 0.04 mg, 0.05 mg, 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg,
0.6 mg, 0.7 mg, 0.8 mg, 0.9 mg, 1.0 mg, 1.5 mg, 2.0 mg, 2.5 mg, 3.0
mg, 3.5 mg, 4.0 mg, 4.5 mg, 5.0 mg, 6.0 mg, 7.0 mg, 8.0 mg, 9.0 mg,
and 10.0 mg; and the VEGF antagonist (e.g., a VEGF trap such as
aflibercept) may be administered to the subject and/or contained in
a co-formulation in an amount selected from the group consisting of
0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.8 mg, 0.9
mg, 1.0 mg, 1.1 mg, 1.2 mg, 1.3 mg, 1.4 mg, 1.5 mg, 1.6 mg, 1.7 mg,
1.8 mg, 1.9 mg, 2.0 mg, 2.1 mg, 2.2 mg, 2.3 mg, 2.4 mg, 2.5 mg, 2.6
mg, 2.7 mg, 2.8 mg, 2.9 mg and 3.0 mg. The
combinations/co-formulations may be administered to a subject
according to any of the administration regimens disclosed elsewhere
herein, including, e.g., twice a week, once every week, once every
2 weeks, once every 3 weeks, once every month, once every 2 months,
once every 3 months, once every 4 months, once every 5 months, once
every 6 months, etc.
Administrative Regimens
[0186] According to certain embodiments of the present invention,
multiple doses of an anti-PD-1 antibody (or a pharmaceutical
composition comprising a combination of an anti-PD-1 antibody and
any of the additional therapeutically active agents mentioned
herein) may be administered to a subject over a defined time
course. The methods according to this aspect of the invention
comprise sequentially administering to a subject multiple doses of
an anti-PD-1 antibody of the invention. As used herein,
"sequentially administering" means that each dose of anti-PD-1
antibody is administered to the subject at a different point in
time, e.g., on different days separated by a predetermined interval
(e.g., hours, days, weeks or months). The present invention
includes methods which comprise sequentially administering to the
patient a single initial dose of an anti-PD-1 antibody, followed by
one or more secondary doses of the anti-PD-1 antibody, and
optionally followed by one or more tertiary doses of the anti-PD-1
antibody. The anti-PD-1 antibody may be administered at a dose
between 0.1 mg/kg to 100 mg/kg.
[0187] The terms "initial dose," "secondary doses," and "tertiary
doses," refer to the temporal sequence of administration of the
anti-PD-1 antibody of the invention. Thus, the "initial dose" is
the dose which is administered at the beginning of the treatment
regimen (also referred to as the "baseline dose"); the "secondary
doses" are the doses which are administered after the initial dose;
and the "tertiary doses" are the doses which are administered after
the secondary doses. The initial, secondary, and tertiary doses may
all contain the same amount of anti-PD-1 antibody, but generally
may differ from one another in terms of frequency of
administration. In certain embodiments, however, the amount of
anti-PD-1 antibody contained in the initial, secondary and/or
tertiary doses varies from one another (e.g., adjusted up or down
as appropriate) during the course of treatment. In certain
embodiments, two or more (e.g., 2, 3, 4, or 5) doses are
administered at the beginning of the treatment regimen as "loading
doses" followed by subsequent doses that are administered on a less
frequent basis (e.g., "maintenance doses").
[0188] In certain exemplary embodiments of the present invention,
each secondary and/or tertiary dose is administered 1 to 26 (e.g.,
1, 11/2, 2, 21/2, 3, 31/2, 4, 41/2, 5, 51/2, 6, 61/2, 7, 71/2, 8,
81/2, 9, 91/2, 10, 101/2, 11, 111/2, 12, 121/2, 13, 131/2, 14,
141/2, 15, 151/2, 16, 161/2, 17, 171/2, 18, 181/2, 19, 191/2, 20,
201/2, 21, 211/2, 22, 221/2, 23, 231/2, 24, 241/2, 25, 251/2, 26,
261/2, or more) weeks after the immediately preceding dose. The
phrase "the immediately preceding dose," as used herein, means, in
a sequence of multiple administrations, the dose of anti-PD-1
antibody which is administered to a patient prior to the
administration of the very next dose in the sequence with no
intervening doses.
[0189] The methods according to this aspect of the invention may
comprise administering to a patient any number of secondary and/or
tertiary doses of an anti-PD-1 antibody. For example, in certain
embodiments, only a single secondary dose is administered to the
patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7,
8, or more) secondary doses are administered to the patient.
Likewise, in certain embodiments, only a single tertiary dose is
administered to the patient. In other embodiments, two or more
(e.g., 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses are
administered to the patient.
[0190] In embodiments involving multiple secondary doses, each
secondary dose may be administered at the same frequency as the
other secondary doses. For example, each secondary dose may be
administered to the patient 1 to 2 weeks or 1 to 2 months after the
immediately preceding dose. Similarly, in embodiments involving
multiple tertiary doses, each tertiary dose may be administered at
the same frequency as the other tertiary doses. For example, each
tertiary dose may be administered to the patient 2 to 12 weeks
after the immediately preceding dose. In certain embodiments of the
invention, the frequency at which the secondary and/or tertiary
doses are administered to a patient can vary over the course of the
treatment regimen. The frequency of administration may also be
adjusted during the course of treatment by a physician depending on
the needs of the individual patient following clinical
examination.
[0191] The present invention includes administration regimens in
which 2 to 6 loading doses are administered to a patient at a first
frequency (e.g., once a week, once every two weeks, once every
three weeks, once a month, once every two months, etc.), followed
by administration of two or more maintenance doses to the patient
on a less frequent basis. For example, according to this aspect of
the invention, if the loading doses are administered at a frequency
of, e.g., once a month (e.g., two, three, four, or more loading
doses administered once a month), then the maintenance doses may be
administered to the patient once every five weeks, once every six
weeks, once every seven weeks, once every eight weeks, once every
ten weeks, once every twelve weeks, etc.).
Diagnostic Uses of the Antibodies
[0192] The anti-PD-1 antibodies of the present invention may be
used to detect and/or measure PD-1 in a sample, e.g., for
diagnostic purposes. Some embodiments contemplate the use of one or
more antibodies of the present invention in assays to detect a
disease or disorder such as cancer, autoimmune disease or chronic
viral infection. Exemplary diagnostic assays for PD-1 may comprise,
e.g., contacting a sample, obtained from a patient, with an
anti-PD-1 antibody of the invention, wherein the anti-PD-1 antibody
is labeled with a detectable label or reporter molecule or used as
a capture ligand to selectively isolate PD-1 from patient samples.
Alternatively, an unlabeled anti-PD-1 antibody can be used in
diagnostic applications in combination with a secondary antibody
which is itself detectably labeled. The detectable label or
reporter molecule can be a radioisotope, such as .sup.3H, .sup.14C,
.sup.32P, .sup.35S, or .sup.125I; a fluorescent or chemiluminescent
moiety such as fluorescein isothiocyanate, or rhodamine; or an
enzyme such as alkaline phosphatase, .beta.-galactosidase,
horseradish peroxidase, or luciferase. Specific exemplary assays
that can be used to detect or measure PD-1 in a sample include
enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA),
and fluorescence-activated cell sorting (FACS).
[0193] Samples that can be used in PD-1 diagnostic assays according
to the present invention include any tissue or fluid sample
obtainable from a patient, which contains detectable quantities of
either PD-1 protein, or fragments thereof, under normal or
pathological conditions. Generally, levels of PD-1 in a particular
sample obtained from a healthy patient (e.g., a patient not
afflicted with cancer or an autoimmune disease) will be measured to
initially establish a baseline, or standard, level of PD-1. This
baseline level of PD-1 can then be compared against the levels of
PD-1 measured in samples obtained from individuals suspected of
having a cancer-related condition, or symptoms associated with such
condition.
[0194] The antibodies specific for PD-1 may contain no additional
labels or moieties, or they may contain an N-terminal or C-terminal
label or moiety. In one embodiment, the label or moiety is biotin.
In a binding assay, the location of a label (if any) may determine
the orientation of the peptide relative to the surface upon which
the peptide is bound. For example, if a surface is coated with
avidin, a peptide containing an N-terminal biotin will be oriented
such that the C-terminal portion of the peptide will be distal to
the surface.
[0195] Aspects of the invention relate to use of the disclosed
antibodies as markers for predicting prognosis of cancer or an
autoimmune disorder in patients. Antibodies of the present
invention may be used in diagnostic assays to evaluate prognosis of
cancer in a patient and to predict survival.
EXAMPLES
[0196] The following examples are put forth so as to provide those
of ordinary skill in the art with a complete disclosure and
description of how to make and use the methods and compositions of
the invention, and are not intended to limit the scope of what the
inventors regard as their invention. Efforts have been made to
ensure accuracy with respect to numbers used (e.g., amounts,
temperature, etc.) but some experimental errors and deviations
should be accounted for. Unless indicated otherwise, parts are
parts by weight, molecular weight is average molecular weight,
temperature is in degrees Centigrade, room temperature is about
25.degree. C., and pressure is at or near atmospheric.
Example 1
Generation of Human Antibodies to PD-1
[0197] Human antibodies to PD-1 were generated using a fragment of
PD-1 that ranges from about amino acids 25-170 of GenBank Accession
NP_005009.2 (SEQ ID NO: 327) with a C93S change. The immunogen was
administered directly, with an adjuvant to stimulate the immune
response, to a VELOCIMMUNE.RTM. mouse comprising DNA encoding human
Immunoglobulin heavy and kappa light chain variable regions. The
antibody immune response was monitored by a PD-1-specific
immunoassay. When a desired immune response was achieved
splenocytes were harvested and fused with mouse myeloma cells to
preserve their viability and form hybridoma cell lines. The
hybridoma cell lines were screened and selected to identify cell
lines that produce PD-1-specific antibodies. Using this technique,
and the immunogen described above, several anti-PD-1 chimeric
antibodies (i.e., antibodies possessing human variable domains and
mouse constant domains) were obtained; exemplary antibodies
generated in this manner were designated as H1M7789N, H1M7799N,
H1M7800N, H2M7780N, H2M7788N, H2M7790N, H2M7791N, H2M7794N,
H2M7795N, H2M7796N, and H2M7798N.
[0198] Anti-PD-1 antibodies were also isolated directly from
antigen-positive B cells without fusion to myeloma cells, as
described in U.S. 2007/0280945A1, herein specifically incorporated
by reference in its entirety. Using this method, several fully
human anti-PD-1 antibodies (i.e., antibodies possessing human
variable domains and human constant domains) were obtained;
exemplary antibodies generated in this manner were designated as
follows: H4H9019P, H4xH9034P2, H4xH9035P2, H4xH9037P2, H4xH9045P2,
H4xH9048P2, H4H9057P2, H4H9068P2, H4xH9119P2, H4xH9120P2,
H4xH9128P2, H4xH9135P2, H4xH9145P2, H4xH8992P, H4xH8999P, and
H4xH9008P.
[0199] The biological properties of the exemplary antibodies
generated in accordance with the methods of this Example are
described in detail in the Examples set forth below.
Example 2
Heavy and Light Chain Variable Region Amino Acid and Nucleotide
Sequences
[0200] Table 1 sets forth the amino acid sequence identifiers of
the heavy and light chain variable regions and CDRs of selected
anti-PD-1 antibodies of the invention. The corresponding nucleic
acid sequence identifiers are set forth in Table 2.
TABLE-US-00001 TABLE 1 Amino Acid Sequence Identifiers Antibody SEQ
ID NOs: Designation HCVR HCDR1 HCDR2 HCDR3 LCVR LCDR1 LCDR2 LCDR3
H1M7789N 2 4 6 8 10 12 14 16 H1M7799N 18 20 22 24 26 28 30 32
H1M7800N 34 36 38 40 42 44 46 48 H2M7780N 50 52 54 56 58 60 62 64
H2M7788N 66 68 70 72 74 76 78 80 H2M7790N 82 84 86 88 90 92 94 96
H2M7791N 98 100 102 104 106 108 110 112 H2M7794N 114 116 118 120
122 124 126 128 H2M7795N 130 132 134 136 138 140 142 144 H2M7796N
146 148 150 152 154 156 158 160 H2M7798N 162 164 166 168 170 172
174 176 H4H9019P 178 180 182 184 186 188 190 192 H4xH9034P2 194 196
198 200 202 204 206 208 H4xH9035P2 210 212 214 216 202 204 206 208
H4xH9037P2 218 220 222 224 202 204 206 208 H4xH9045P2 226 228 230
232 202 204 206 208 H4xH9048P2 234 236 238 240 202 204 206 208
H4H9057P2 242 244 246 248 202 204 206 208 H4H9068P2 250 252 254 256
202 204 206 208 H4xH9119P2 258 260 262 264 202 204 206 208
H4xH9120P2 266 268 270 272 202 204 206 208 H4xH9128P2 274 276 278
280 202 204 206 208 H4xH9135P2 282 284 286 288 202 204 206 208
H4xH9145P2 290 292 294 296 202 204 206 208 H4xH8992P 298 300 302
304 186 188 190 192 H4xH8999P 306 308 310 312 186 188 190 192
H4xH9008P 314 316 318 320 186 188 190 192
TABLE-US-00002 TABLE 2 Nucleic Acid Sequence Identifiers Antibody
SEQ ID NOs: Designation HCVR HCDR1 HCDR2 HCDR3 LCVR LCDR1 LCDR2
LCDR3 H1M7789N 1 3 5 7 9 11 13 15 H1M7799N 17 19 21 23 25 27 29 31
H1M7800N 33 35 37 39 41 43 45 47 H2M7780N 49 51 53 55 57 59 61 63
H2M7788N 65 67 69 71 73 75 77 79 H2M7790N 81 83 85 87 89 91 93 95
H2M7791N 97 99 101 103 105 107 109 111 H2M7794N 113 115 117 119 121
123 125 127 H2M7795N 129 131 133 135 137 139 141 143 H2M7796N 145
147 149 151 153 155 157 159 H2M7798N 161 163 165 167 169 171 173
175 H4H9019P 177 179 181 183 185 187 189 191 H4xH9034P2 193 195 197
199 201 203 205 207 H4xH9035P2 209 211 213 215 201 203 205 207
H4xH9037P2 217 219 221 223 201 203 205 207 H4xH9045P2 225 227 229
231 201 203 205 207 H4xH9048P2 233 235 237 239 201 203 205 207
H4H9057P2 241 243 245 247 201 203 205 207 H4H9068P2 249 251 253 255
201 203 205 207 H4xH9119P2 257 259 261 263 201 203 205 207
H4xH9120P2 265 267 269 271 201 203 205 207 H4xH9128P2 273 275 277
279 201 203 205 207 H4xH9135P2 281 283 285 287 201 203 205 207
H4xH9145P2 289 291 293 295 201 203 205 207 H4xH8992P 297 299 301
303 185 187 189 191 H4xH8999P 305 307 309 311 185 187 189 191
H4xH9008P 313 315 317 319 185 187 189 191
[0201] Antibodies are typically referred to herein according to the
following nomenclature: Fc prefix (e.g. "H4xH," "H1M," "H2M,"
etc.), followed by a numerical identifier (e.g. "7789," "7799,"
etc., as shown in Table 1), followed by a "P," "P2," "N," or "B"
suffix. Thus, according to this nomenclature, an antibody may be
referred to herein as, e.g., "H1H7789N," "H1M7799N," "H2M7780N,"
etc. The H4xH, HIM, H2M and H2aM prefixes on the antibody
designations used herein indicate the particular Fc region isotype
of the antibody. For example, an "H4xH" antibody has a human IgG4
Fc with 2 or more amino acid changes as disclosed in US20100331527,
an "H1M" antibody has a mouse IgG1 Fc, and an "H2M" antibody has a
mouse IgG2 Fc (a or b isotype) (all variable regions are fully
human as denoted by the first `H` in the antibody designation). As
will be appreciated by a person of ordinary skill in the art, an
antibody having a particular Fc isotype can be converted to an
antibody with a different Fc isotype (e.g., an antibody with a
mouse IgG1 Fc can be converted to an antibody with a human IgG4,
etc.), but in any event, the variable domains (including the
CDRs)--which are indicated by the numerical identifiers shown in
Table 1--will remain the same, and the binding properties to
antigen are expected to be identical or substantially similar
regardless of the nature of the Fc domain.
[0202] In certain embodiments, selected antibodies with a mouse
IgG1 Fc were converted to antibodies with human IgG4 Fc. In one
embodiment, the IgG4 Fc domain comprises a serine to proline
mutation in the hinge region (S108P) to promote dimer
stabilization. Table 3 sets forth the amino acid sequence
identifiers of heavy chain and light chain sequences of selected
anti-PD-1 antibodies with human IgG4 Fc.
TABLE-US-00003 TABLE 3 Antibody SEQ ID NOs: Designation Heavy Chain
Light Chain H4H7798N 330 331 H4H7795N2 332 333 H4H9008P 334 335
H4H9048P2 336 337
[0203] Each heavy chain sequence in Table 3 comprised a variable
region (V.sub.H or HCVR; comprising HCDR1, HCDR2 and HCDR3) and a
constant region (comprising C.sub.H1, C.sub.H2 and C.sub.H3
domains). Each light chain sequence in Table 3 comprised a variable
region (V.sub.L or LCVR; comprising LCDR1, LCDR2 and LCDR3) and a
constant region (C.sub.L). SEQ ID NO: 330 comprised a HCVR
comprising amino acids 1-117 and a constant region comprising amino
acids 118-444. SEQ ID NO: 331 comprised a LCVR comprising amino
acids 1-107 and a constant region comprising amino acids 108-214.
SEQ ID NO: 332 comprised a HCVR comprising amino acids 1-122 and a
constant region comprising amino acids 123-449. SEQ ID NO: 333
comprised a LCVR comprising amino acids 1-107 and a constant region
comprising amino acids 108-214. SEQ ID NO: 334 comprised a HCVR
comprising amino acids 1-119 and a constant region comprising amino
acids 120-446. SEQ ID NO: 335 comprised a LCVR comprising amino
acids 1-108 and a constant region comprising amino acids 109-215.
SEQ ID NO: 336 comprised a HCVR comprising amino acids 1-121 and a
constant region comprising amino acids 122-448. SEQ ID NO: 337
comprised a LCVR comprising amino acids 1-108 and a constant region
comprising amino acids 109-215.
Example 3
Antibody Binding to PD-1 as Determined by Surface Plasmon
Resonance
[0204] Binding association and dissociation rate constants (k.sub.a
and k.sub.d, respectively), equilibrium dissociation constants and
dissociation half-lives (K.sub.D and t.sub.1/2, respectively) for
antigen binding to purified anti-PD1 antibodies were determined
using a real-time surface plasmon resonance biosensor assay on a
Biacore 4000 or Biacore T200 instrument. The Biacore sensor surface
was derivatized with either a polyclonal rabbit anti-mouse antibody
(GE, #BR-1008-38) or with a monoclonal mouse anti-human Fc antibody
(GE, #BR-1008-39) to capture approximately 100-900 RUs of anti-PD-1
monoclonal antibodies, expressed with either a mouse Fc or a human
Fc, respectively. The PD-1 reagents tested for binding to the
anti-PD-1 antibodies included recombinant human PD-1 expressed with
a C-terminal myc-myc-hexahistidine tag (hPD-1-MMH; SEQ ID NO: 321),
recombinant cynomolgus monkey PD-1 expressed with a C-terminal
myc-myc-hexahistidine tag (MfPD-1-MMH; SEQ ID NO: 322), recombinant
human PD-1 dimer expressed with either a C-terminal mouse IgG2a Fc
tag (hPD-1-mFc; SEQ ID NO: 323) or with a C-terminal human IgG1 Fc
(hPD1-hFc; SEQ ID NO: 324), and monkey PD-1 with mFc (SEQ ID NO:
329). Different concentrations of PD-1 reagents ranging from 200 nM
to 3.7 nM were injected over the anti-PD-1 monoclonal antibody
captured surface at a flow rate of 30 .mu.L/min on Biacore 4000 or
at 50 .mu.L/min on Biacore T200. The binding of the PD-1 reagents
to captured monoclonal antibodies was monitored for 3 to 5 minutes
while their dissociation from the antibodies was monitored for 7 to
10 minutes in HBST running buffer (0.01 M HEPES pH 7.4, 0.15 M
NaCl, 3 mM EDTA, 0.05% v/v Surfactant P20). Experiments were
performed at 25.degree. C. and 37.degree. C. Kinetic association
(k.sub.a) and dissociation (k.sub.d) rate constants were determined
by processing and fitting the data to a 1:1 binding model using
Scrubber 2.0c curve fitting software. Binding dissociation
equilibrium constants (K.sub.D) and dissociative half-lives
(t.sub.1/2) were then calculated from the kinetic rate constants
as: K.sub.D (M)=k.sub.d/k.sub.a and t.sub.1/2
(min)=[In2/(60*k.sub.d)]. Binding kinetics parameters for different
anti-PD-1 monoclonal antibodies binding to different PD-1 reagents
at 25.degree. C. and 37.degree. C. are tabulated in Tables
4-11.
TABLE-US-00004 TABLE 4 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to human PD-1-MMH at 25.degree. C.
k.sub.a k.sub.d K.sub.D t1/2 Antibody (1/Ms) (1/s) (M) (min)
H2aM7780N 9.32E+03 3.59E-04 3.85E-08 32 H2aM7788N 1.97E+04 3.88E-04
1.96E-08 30 H1M7789N 2.53E+04 5.31E-05 2.10E-09 218 H2aM7790N
4.63E+04 8.23E-04 1.78E-08 14 H2aM7791N 3.01E+04 7.06E-04 2.34E-08
16 H2aM7794N 5.50E+04 2.12E-03 3.80E-08 5.4 H2aM7795N 4.91E+04
1.15E-03 2.35E-08 10 H2aM7796N 6.73E+03 1.93E-03 2.86E-07 6.0
H2aM7798N 1.32E+05 3.06E-04 2.31E-09 38 H1M7799N 5.04E+04 1.23E-02
2.44E-07 0.9 H1M7800N 5.88E+04 9.47E-03 1.61E-07 1.2 H4H9019P
2.05E+04 8.08E-04 3.94E-08 14 H4xH9034P 1.02E+05 1.49E-03 1.45E-08
7.8 H4xH9035P 1.03E+05 4.75E-04 4.62E-09 24 H4xH9037P 7.32E+04
7.95E-04 1.09E-08 15 H4xH9045P 5.40E+04 4.03E-03 7.46E-08 2.9
H4xH9048P2 1.37E+05 1.23E-03 8.95E-09 9.4 H4H9057P2 4.60E+04
1.34E-02 2.91E-07 0.9 H4H9068P2 NB* NB* NB* NB* H4xH9119P2 7.84E+04
1.22E-03 1.56E-08 9.5 H4xH9120P2 3.32E+04 9.98E-04 3.01E-08 12
H4xH9128P2 4.95E+04 7.19E-04 1.45E-08 16 H4xH9135P2 1.17E+05
1.20E-03 1.02E-08 10 H4xH9145P2 3.47E+04 1.34E-03 3.85E-08 8.6
H4xH8992P 1.50E+05 2.13E-02 1.41E-07 0.5 H4xH8999P 2.83E+05
1.23E-03 4.33E-09 9.4 H4xH9008P 4.29E+04 1.33E-03 3.10E-08 8.7
H4H7795N2 6.35E+04 1.48E-03 2.33E-08 8 H4H7798N 1.47E+05 4.43E-04
3.01E-09 26 *NB indicates that under the experimental conditions,
PD-1 reagent did not bind to the captured anti-PD-1 monoclonal
antibody
TABLE-US-00005 TABLE 5 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to human PD-1-MMH at 37.degree. C.
k.sub.a k.sub.d K.sub.D t1/2 Antibody (1/Ms) (1/s) (M) (min)
H2aM7780N 2.72E+04 1.52E-03 5.58E-08 7.6 H2aM7788N 2.88E+04
1.49E-03 5.19E-08 7.7 H1M7789N 4.53E+04 2.95E-04 6.52E-09 39
H2aM7790N 6.13E+04 5.20E-03 8.49E-08 2.2 H2aM7791N 4.18E+04
2.24E-03 5.35E-08 5.2 H2aM7794N 1.20E+05 7.92E-03 6.61E-08 1.5
H2aM7795N 6.75E+04 4.58E-03 6.78E-08 2.5 H2aM7796N 1.09E+04
1.65E-02 1.51E-06 0.7 H2aM7798N 1.73E+05 6.56E-04 3.79E-09 18
H1M7799N 7.94E+04 4.25E-02 5.36E-07 0.3 H1M7800N 7.83E+04 3.99E-02
5.10E-07 0.3 H4H9019P 1.20E+04 5.44E-03 4.53E-07 2.1 H4xH9034P
2.79E+05 1.12E-02 4.02E-08 1.0 H4xH9035P 2.98E+05 4.26E-03 1.43E-08
2.7 H4xH9037P 2.26E+05 6.68E-03 2.95E-08 1.7 H4xH9045P 8.04E+04
5.32E-02 6.62E-07 0.2 H4xH9048P2 3.70E+05 8.60E-03 2.32E-08 1.3
H4H9057P2 NB* NB* NB* NB* H4H9068P2 NB* NB* NB* NB* H4xH9119P2
2.40E+05 1.04E-02 4.35E-08 1.1 H4xH9120P2 6.88E+04 7.01E-03
1.02E-07 1.6 H4xH9128P2 1.04E+05 4.36E-03 4.20E-08 2.6 H4xH9135P2
4.18E+05 1.11E-02 2.66E-08 1.0 H4xH9145P2 1.31E+05 1.23E-02
9.40E-08 0.9 H4xH8992P IC* IC* IC* IC* H4xH8999P 5.99E+05 9.42E-03
1.57E-08 1.2 H4xH9008P 1.29E+05 8.09E-03 6.26E-08 1.4 H4H7795N2
6.41E+04 6.64E-03 1.04E-07 1.7 H4H7798N 2.27E+05 1.70E-03 7.48E-09
7 *NB indicates that under the experimental conditions, PD-1
reagent did not bind to the captured anti-PD-1 monoclonal antibody.
IC indicates that under the experimental conditions, PD-1 binding
is inconclusive.
TABLE-US-00006 TABLE 6 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to human PD-1 dimer (human PD-1-mFc
or human PD-1-hFc) at 25.degree. C. k.sub.a k.sub.d K.sub.D t1/2
Antibody (1/Ms) (1/s) (M) (min) H2aM7780N 4.21E+04 9.94E-06
2.36E-10 1162 H2aM7788N 8.94E+04 2.82E-05 3.15E-10 410 H1M7789N
3.91E+04 4.31E-05 1.10E-09 268 H2aM7790N 1.86E+05 3.02E-05 1.62E-10
383 H2aM7791N 4.05E+04 1.01E-04 2.49E-09 114 H2aM7794N 1.79E+05
1.06E-04 5.93E-10 109 H2aM7795N 1.38E+05 3.14E-05 2.27E-10 368
H2aM7796N 2.61E+04 8.67E-05 3.32E-09 133 H2aM7798N 3.50E+05
2.29E-05 6.55E-11 505 H1M7799N 2.38E+05 8.55E-05 3.60E-10 135
H1M7800N 1.52E+05 7.72E-05 5.09E-10 150 H4H9019P 4.38E+04 8.61E-05
1.97E-09 134 H4xH9034P 2.15E+05 1.51E-04 7.01E-10 77 H4xH9035P
2.01E+05 1.03E-04 5.13E-10 112 H4xH9037P 1.50E+05 1.29E-04 8.62E-10
89 H4xH9045P 9.13E+04 1.60E-04 1.75E-09 72 H4xH9048P2 2.36E+05
1.88E-04 7.98E-10 61 H4H9057P2 1.01E+05 1.77E-04 1.75E-09 65
H4H9068P2 4.72E+04 2.80E-03 5.94E-08 4 H4xH9119P2 1.63E+05 1.62E-04
9.92E-10 71 H4xH9120P2 6.52E+04 1.19E-04 1.82E-09 97 H4xH9128P2
8.37E+04 1.33E-04 1.59E-09 87 H4xH9135P2 2.12E+05 1.38E-04 6.51E-10
84 H4xH9145P2 6.58E+04 1.58E-04 2.40E-09 73 H4xH8992P 2.35E+05
1.60E-04 6.80E-10 72 H4xH8999P 5.55E+05 1.20E-04 2.17E-10 96
H4xH9008P 3.52E+04 2.80E-05 7.96E-10 412 H4H7795N2 1.50E+05
9.25E-05 6.15E-10 125 H4H7798N 4.41E+05 5.40E-05 1.22E-10 214
TABLE-US-00007 TABLE 7 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to human PD-1 dimer (human PD-1-mFc
or human PD-1-hFc) at 37.degree. C. k.sub.a k.sub.d K.sub.D t1/2
Antibody (1/Ms) (1/s) (M) (min) H2aM7780N 9.94E+04 2.29E-05
2.30E-10 505 H2aM7788N 1.31E+05 2.13E-05 1.63E-10 542 H1M7789N
1.09E+05 .ltoreq.1.0E-05 .ltoreq.9.17E-11 .gtoreq.1155 H2aM7790N
2.01E+05 8.49E-05 4.22E-10 136 H2aM7791N 4.98E+04 1.79E-04 3.59E-09
65 H2aM7794N 4.68E+05 2.11E-04 4.52E-10 55 H2aM7795N 1.65E+05
6.13E-05 3.71E-10 188 H2aM7796N 2.21E+04 4.34E-04 1.96E-08 27
H2aM7798N 4.90E+05 1.40E-05 2.80E-11 825 H1M7799N 4.41E+05 1.81E-04
4.11E-10 64 H1M7800N 4.00E+05 1.81E-04 4.50E-10 64 H4H9019P
7.17E+04 1.95E-04 2.71E-09 59 H4xH9034P 3.02E+05 6.30E-04 2.09E-09
18 H4xH9035P 3.16E+05 5.54E-04 1.75E-09 21 H4xH9037P 2.63E+05
9.21E-04 3.50E-09 13 H4xH9045P 2.14E+05 1.10E-03 5.13E-09 11
H4xH9048P2 3.61E+05 1.10E-03 3.05E-09 10 H4H9057P2 2.33E+05
2.11E-03 9.07E-09 5 H4H9068P2 9.69E+04 1.20E-02 1.24E-07 1
H4xH9119P2 2.40E+05 9.09E-04 3.80E-09 13 H4xH9120P2 8.08E+04
4.82E-04 5.96E-09 24 H4xH9128P2 1.86E+05 6.86E-04 3.68E-09 17
H4xH9135P2 3.10E+05 7.02E-04 2.27E-09 16 H4xH9145P2 1.60E+05
5.71E-04 3.58E-09 20 H4xH8992P 3.49E+05 1.02E-03 2.91E-09 11
H4xH8999P 7.57E+05 4.51E-04 5.96E-10 26 H4xH9008P 5.52E+04
.ltoreq.1.0E-05 .ltoreq.1.81E-10 .gtoreq.1155 H4H7795N2 1.60E+05
2.64E-04 1.65E-09 44 H4H7798N 6.60E+05 1.15E-04 1.75E-10 100
TABLE-US-00008 TABLE 8 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to MfPD-1-MMH at 25.degree. C.
k.sub.a k.sub.d K.sub.D t1/2 Antibody (1/Ms) (1/s) (M) (min)
H2aM7780N 1.00E+04 3.15E-04 3.15E-08 37 H2aM7788N 8.63E+03 6.62E-04
7.66E-08 17 H1M7789N 1.55E+04 1.23E-04 7.89E-09 94 H2aM7790N
3.11E+04 9.37E-04 3.01E-08 12 H2aM7791N 1.61E+04 5.53E-04 3.44E-08
21 H2aM7794N 3.60E+04 5.99E-03 1.66E-07 1.9 H2aM7795N 4.44E+04
8.89E-04 2.01E-08 13 H2aM7796N NB* NB* NB* NB* H2aM7798N 8.72E+04
3.93E-04 4.50E-09 29 H1M7799N 5.78E+04 1.30E-02 2.24E-07 0.9
H1M7800N 5.89E+04 1.04E-02 1.76E-07 1.1 H4H9019P 1.94E+04 8.33E-04
4.29E-08 14 H4xH9034P 9.61E+04 2.69E-03 2.80E-08 4.3 H4xH9035P
9.36E+04 4.34E-04 4.64E-09 27 H4xH9037P 6.99E+04 9.15E-04 1.31E-08
13 H4xH9045P 6.25E+04 7.05E-03 1.13E-07 1.6 H4xH9048P2 1.28E+05
8.97E-04 7.00E-09 13 H4H9057P2 3.46E+04 1.91E-02 5.51E-07 0.6
H4H9068P2 NB* NB* NB* NB* H4xH9119P2 7.50E+04 1.66E-03 2.22E-08 6.9
H4xH9120P2 3.17E+04 1.08E-03 3.41E-08 11 H4xH9128P2 3.68E+04
6.49E-04 1.77E-08 18 H4xH9135P2 1.24E+05 1.31E-03 1.06E-08 8.8
H4xH9145P2 2.86E+04 1.24E-03 4.31E-08 9.3 H4xH8992P 1.88E+05
3.76E-02 2.00E-07 0.3 H4xH8999P 4.29E+05 1.33E-03 3.09E-09 8.7
H4xH9008P 1.05E+05 2.49E-03 2.38E-08 4.6 H4H7795N2 6.59E+04
1.48E-03 2.24E-08 8 H4H7798N 1.43E+05 5.51E-04 3.86E-09 21 *NB
indicates that under the experimental conditions, PD-1 reagent did
not bind to the captured anti-PD-1 monoclonal antibody
TABLE-US-00009 TABLE 9 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to MfPD-1-MMH at 37.degree. C.
k.sub.a k.sub.d K.sub.D t1/2 Antibody (1/Ms) (1/s) (M) (min)
H2aM7780N 2.29E+04 1.38E-03 6.05E-08 8.3 H2aM7788N 1.88E+04
3.28E-03 1.74E-07 3.5 H1M7789N 4.79E+04 4.08E-04 8.50E-09 28
H2aM7790N 2.55E+04 6.93E-03 2.71E-07 1.7 H2aM7791N 3.79E+04
1.91E-03 5.05E-08 6.0 H2aM7794N 6.66E+04 2.01E-02 3.02E-07 0.6
H2aM7795N 6.47E+04 3.89E-03 6.02E-08 3.0 H2aM7796N NB* NB* NB* NB*
H2aM7798N 1.42E+05 9.93E-04 7.00E-09 12 H1M7799N 8.80E+04 4.67E-02
5.30E-07 0.2 H1M7800N 8.40E+04 4.43E-02 5.27E-07 0.3 H4H9019P
2.14E+04 7.63E-03 3.56E-07 1.5 H4xH9034P 2.83E+05 2.47E-02 8.73E-08
0.5 H4xH9035P 3.06E+05 4.29E-03 1.40E-08 2.7 H4xH9037P 2.22E+05
8.80E-03 3.97E-08 1.3 H4xH9045P 1.40E+04 1.05E-01 7.54E-06 0.1
H4xH9048P2 4.15E+05 6.97E-03 1.68E-08 1.7 H4H9057P2 NB* NB* NB* NB*
H4H9068P2 NB* NB* NB* NB* H4xH9119P2 2.40E+05 1.23E-02 5.14E-08 0.9
H4xH9120P2 6.98E+04 7.48E-03 1.07E-07 1.5 H4xH9128P2 9.06E+04
4.18E-03 4.61E-08 2.8 H4xH9135P2 4.62E+05 1.34E-02 2.89E-08 0.9
H4xH9145P2 1.71E+05 1.43E-02 8.37E-08 0.8 H4xH8992P IC* IC* IC* IC*
H4xH8999P 9.83E+05 9.26E-03 9.41E-09 1.2 H4xH9008P 5.86E+05
1.38E-02 2.35E-08 0.8 H4H7795N2 7.80E+04 6.89E-03 8.83E-08 1.7
H4H7798N 2.13E+05 2.23E-3 1.05E-08 5 *NB indicates that under the
experimental conditions, PD-1 reagent did not bind to the captured
anti-PD-1 monoclonal antibody. IC indicates that under the
experimental conditions, PD-1 binding is inconclusive.
TABLE-US-00010 TABLE 10 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to monkey PD-1 dimer (monkey
PD-1-mFc) at 25.degree. C. 100 nM Amount of Monkey mAb PD-1-mFc
Captured Bound k.sub.a k.sub.d K.sub.D t1/2 Antibody (RU) (RU)
(1/Ms) (1/s) (M) (min) H4H9019P 116 31 4.55E+04 8.96E-05 1.97E-09
129 H4xH9034P 215 95 2.03E+05 1.66E-04 8.18E-10 70 H4xH9035P 153 78
2.16E+05 9.96E-05 4.60E-10 116 H4xH9037P 137 58 1.50E+05 1.37E-04
9.12E-10 84 H4xH9045P 202 78 9.78E+04 1.68E-04 1.72E-09 69
H4xH9048P2 227 115 2.43E+05 1.84E-04 7.54E-10 63 H4H9057P2 196 75
1.02E+05 3.03E-04 2.98E-09 38 H4H9068P2 178 17 5.70E+04 3.09E-03
5.42E-08 4 H4xH9119P2 209 83 1.63E+05 1.72E-04 1.05E-09 67
H4xH9120P2 195 52 5.84E+04 1.12E-04 1.91E-09 104 H4xH9128P2 175 64
7.87E+04 1.24E-04 1.57E-09 94 H4xH9135P2 150 74 2.38E+05 1.43E-04
6.02E-10 81 H4xH9145P2 304 84 7.24E+04 1.50E-04 2.08E-09 77
H4xH8992P 260 122 2.03E+05 2.51E-04 1.24E-09 46 H4xH8999P 217 126
5.50E+05 1.15E-04 2.10E-10 100 H4xH9008P 248 93 1.20E+05 5.77E-05
4.80E-10 200 H4H7795N2 204 60 1.60E+05 9.92E-05 6.21E-10 116
H4H7798N 223 93 4.49E+05 6.14E-05 1.37E-10 188
TABLE-US-00011 TABLE 11 Binding Kinetics parameters of anti-PD-1
monoclonal antibodies binding to monkey PD-1 dimer (monkey
PD-1-mFc) at 37.degree. C. 100 nM Amount of Monkey mAb PD-1-mFc
Captured Bound k.sub.a k.sub.d K.sub.D t1/2 Antibody (RU) (RU)
(1/Ms) (1/s) (M) (min) H4H9019P 89 36 8.16E+04 2.59E-04 3.17E-09 45
H4xH9034P 184 81 3.07E+05 7.49E-04 2.44E-09 15 H4xH9035P 88 40
3.67E+05 6.23E-04 1.70E-09 19 H4xH9037P 55 24 2.80E+05 8.97E-04
3.21E-09 13 H4xH9045P 161 65 2.41E+05 1.36E-03 5.66E-09 8
H4xH9048P2 184 84 4.94E+05 1.13E-03 2.29E-09 10 H4H9057P2 105 28
1.61E+05 4.77E-03 2.96E-08 2.4 H4H9068P2 90 6 1.21E+05 1.05E-02
8.63E-08 1.1 H4xH9119P2 98 40 2.79E+05 8.85E-04 3.17E-09 13
H4xH9120P2 141 46 8.29E+04 5.02E-04 6.06E-09 23 H4xH9128P2 148 60
1.87E+05 8.16E-04 4.36E-09 14 H4xH9135P2 106 52 3.42E+05 7.94E-04
2.32E-09 15 H4xH9145P2 284 94 1.51E+05 6.09E-04 4.04E-09 19
H4xH8992P 206 86 3.50E+05 1.53E-03 4.38E-09 `8 H4xH8999P 160 83
7.30E+05 5.10E-04 7.00E-10 23 H4xH9008P 216 98 2.04E+05 1.00E-05*
4.90E-11* 1155* H4H7795N2 164 47 1.70E+05 2.90E-04 1.71E-09 40
H4H7798N 203 88 6.30E+05 1.27E-04 2.02E-10 91 *indicates that under
the current experimental conditions, no dissociation of PD-1
reagent was observed and the value of kd was manually fixed at
1.00E-05
[0205] As shown in Table 4, at 25.degree. C., 28 of the 29
anti-PD-1 antibodies of the invention bound to hPD-1-MMH with
K.sub.D values ranging from 2.1 nM to 291 nM. One antibody,
H4H9068P2, did not demonstrate any measurable binding to hPD-1-MMH
at 25.degree. C. As shown in Table 5, at 37.degree. C., 26 of the
29 anti-PD-1 antibodies of the invention bound to hPD-1-MMH with
K.sub.D values ranging from 3.79 nM to 1.51 .mu.M. Three antibodies
of the invention did not demonstrate any conclusive binding to
hPD-1-MMH at 37.degree. C. As shown in Table 6, at 25.degree. C.,
all 29 anti-PD-1 antibodies of the invention bound to hPD-1 dimer
proteins with K.sub.D values ranging from 65.5 .mu.M to 59.4 nM. As
shown in Table 7, at 37.degree. C., all 27 anti-PD-1 antibodies of
the invention bound to hPD-1 dimer proteins with K.sub.D values
ranging from 3.09 .mu.M to 551 nM. As shown in Table 8, at
25.degree. C., 27 of the 29 anti-PD-1 antibodies of the invention
bound to MfPD-1-MMH with K.sub.D values ranging from 3.09 nM to 551
nM. Two antibodies of the invention did not demonstrate any
conclusive binding to MfPD-1-MMH at 25.degree. C. As shown in Table
9, at 37.degree. C., 25 of the 29 anti-PD-1 antibodies of the
invention bound to MfPD-1-MMH with K.sub.D values ranging from 7.00
nM to 7.54 .mu.M. Four antibodies of the invention did not
demonstrate any conclusive binding to MfPD-1-MMH at 37.degree. C.
As shown in Table 10, at 25.degree. C., all 18 of the tested
anti-PD-1 antibodies of the invention bound to MfPD-1 dimer with
K.sub.D values ranging from 137 .mu.M to 54.2 nM. As shown in Table
11, at 37.degree. C., all 18 of the tested anti-PD-1 antibodies of
the invention bound to MfPD-1 dimer with K.sub.D values ranging
from less than 49 .mu.M to 86.3 nM.
Example 4
Blocking of PD-1 Binding to PD-L1 as Determined by ELISA
[0206] The ability of anti-PD-1 antibodies to block human PD-1
binding to its ligand, the PD-L1 receptor, was measured using three
competition sandwich ELISA formats. Dimeric human PD-L1 proteins,
comprised of a portion of the human PD-L1 extracellular domain
expressed with either a C-terminal human Fc tag (hPD-L1-hFc; SEQ
ID: 325) or a C-terminal mouse Fc tag (hPD-L1-mFc; SEQ ID: 326), or
dimeric human PD-L2, comprised of the human PD-L2 extracellular
region produced with a C-terminal human Fc tag (hPD-L2-hFc; R&D
Systems, #1224-PL) were separately coated at a concentration of 2
.mu.g/mL in PBS on a 96-well microtiter plate overnight at
4.degree. C. Nonspecific binding sites were subsequently blocked
using a 0.5% (w/v) solution of BSA in PBS. In a first competition
format, a constant concentration of 1.5 nM of a dimeric human PD-1
protein, comprised of the human PD-1 extracellular domain expressed
with a C-terminal mouse Fc tag (hPD-1-mFc; SEQ ID: 323) was added
to serial dilutions of anti-PD-1 antibodies or isotype control
antibodies so that the final concentrations of antibodies ranged
from 0 to 200 nM. In a second competition format, a constant
concentration of 200 pM of dimeric biotinylated human PD-1 protein,
comprised of the human PD-1 extracellular domain that was expressed
with a C-terminal human Fc tag (biot-hPD-1-hFc; SEQ ID: 323), was
similarly added to serial dilutions of anti-PD-1 antibodies or an
isotype control at final antibody concentrations ranging from 0 to
50 nM. In a third competition format, a constant concentration of
100 pM of dimeric hPD-1-mFc protein was similarly added to serial
dilutions of anti-PD-1 antibodies or an isotype control at final
antibody concentrations ranging from 0 to 100 nM. These
antibody-protein complexes were then incubated for 1 hour at room
temperature (RT). Antibody-protein complexes with 1.5 nM constant
hPD-1-mFc were transferred to microtiter plates coated with
hPD-L1-hFc, antibody-protein complexes with 200 pM constant
biot-hPD-1-hFc were transferred to hPD-L1-mFc coated plates, and
antibody-protein complexes with 100 pM constant hPD-1-mFc were
transferred to microtiter plates coated with hPD-L2-hFc. After
incubating for 1 hour at RT, the wells were washed, and plate-bound
hPD-1-mFc was detected with an anti-mFc polyclonal antibody
conjugated with horseradish peroxidase (HRP) (Jackson
ImmunoResearch Inc., #115-035-164), and plate-bound biot-hPD-1-hFc
was detected with streptavidin conjugated with HRP (Thermo
Scientific, #N200). Samples were developed with a TMB solution (BD
Biosciences, #51-2606KC and #51-2607KC) to produce a colorimetric
reaction and then color development was stabilized by addition of
1M sulfuric acid before measuring absorbance at 450 nm on a Victor
X5 plate reader. Data analysis was performed using a sigmoidal
dose-response model within Prism.TM. software (GraphPad). The
calculated IC.sub.50 value, defined as the concentration of
antibody required to reduce 50% of human PD-1 binding to human
PD-L1 or PD-L2, was used as an indicator of blocking potency.
Percent maximum blockade was calculated as a measure of the ability
of the antibodies to completely block binding of human PD-1 to
human PD-L1 or PD-L2 on the plate as determined from the dose
curve. This percent maximum blockade was calculated by subtracting
from 100% the ratio of the reduction in signal observed in the
presence of the highest tested concentration for each antibody
relative to the difference between the signal observed for a sample
of human PD-1 containing no anti-PD-1 antibody (0% blocking) and
the background signal from HRP-conjugated secondary antibody alone
(100% blocking).
[0207] Percent maximum blockade and the calculated IC.sub.50 values
for antibodies blocking greater than 35% of the hPD-1 binding
signal are shown in Tables 12-14. Antibodies that showed a decrease
in the hPD-1 binding signal of 35% or less were defined as
non-blockers. Antibodies that showed an increase of 35% or more in
the binding signal of human PD-1 were characterized as
non-blocker/enhancers. The theoretical assay bottom, defined as the
minimum antibody concentration theoretically needed to occupy 50%
binding sites of human PD-1 in the assay, is 0.75 nM for the format
using 1.5 nM constant hPD-1-mFc, 100 pM for the format using 200 pM
constant biot-hPD-1-hFc, and 50 pM for the format using 100 pM
constant hPD-1-mFc, indicating that lower calculated IC.sub.50
values may not represent quantitative protein-antibody site
binding. For this reason, antibodies with calculated IC.sub.50
values less than 0.75 nM in the assay with hPD-1-mFc constant and
hPD-L1 coat, less than 100 pM in the assay with biot-hPD-1-hFc
constant and hPD-L1 coat, and less than 50 pM in the assay with
hPD-1-mFc constant and hPD-L2 coat are reported in Tables 12-14 as
<7.5E-10M, <1.0E-10M and <5.0E-11M, respectively.
TABLE-US-00012 TABLE 12 ELISA blocking of human PD-1 binding to
human PD-L1 by anti-PD-1 antibodies Blocking 1.5 nM 200 nM Antibody
of hPD-1-mFc blocking 1.5 nM binding to hPD-1-mFc binding
hPD-L1-hFc, to hPD-L1-hFc, Antibody IC.sub.50 (M) % blocking
H4H9019P 1.3E-09 98 H4xH9034P 5.1E-10* 98 H4xH9045P 2.8E-10* 98
H4xH9048P2 3.3E-09 67 H4xH9120P2 1.0E-09 98 H4xH9128P2 6.4E-10* 98
H4xH9035P 6.2E-10* 99 H4xH9135P2 1.1E-09 97 H4xH9145P2 9.3E-10 90
H4xH9119P2 2.0E-10* 78 H4H9057P2 1.9E-10* 98 H4H9068P2
NBI/Enchancer -142 H4xH9037P 8.9E-10 100 H2aM7780N 6.9E-10* 94
H2aM7788N 2.2E-10* 74 H1M7789N NBI/Enchancer -170 H2aM7790N 1.5E-09
74 H2aM7791N NBI/Enchancer -154 H2aM7794N 1.1E-09 95 H2aM7795N2
8.6E-10 93 H2aM7796N NBI -20 H2aM7798N 6.8E-10* 93 H1M7799N
2.2E-10* 82 H1M7800N 6.0E-10* 83 H4xH8992P 1.3E-09 93 H4xH8999P
1.3E-09 88 H4xH9008P 2.4E-09 88 Isotype control 1 NBI -3 Isotype
control 2 NBI -34 Isotype control 2 NBI -7 Isotype control 2 NBI
-16 Assay theoretical bottom: for blocking ELISA with hPD-1-mFc
constant and hPD-L1 coat is 7.5E-10M *Below theoretical bottom of
the assay; NT - not tested; NBI - non-blocker; NBI/Enhancer -
non-blocker/enhancer; IC - inconclusive
TABLE-US-00013 TABLE 13 ELISA blocking of biotinylated human PD-1
binding to human PD-L1 by anti-PD-1 antibodies 50 nM Antibody
blocking Blocking 200 pM biot- 200 pM biot-hPD-1-hFc hPD-1-hFc
binding to binding to hPD-L1-mFc, Antibody hPD-L1-mFc, IC.sub.50
(M) % blocking H4H9019P 6.4E-10 97 H4xH9034P 6.6E-11* 96 H4xH9045P
1.3E-10 95 H4xH9048P2 IC 76 H4xH9120P2 3.9E-10 96 H4xH9128P2
1.9E-10 97 H4xH9035P 8.0E-11* 95 H4xH9135P2 1.5E-10 96 H4xH9145P2
3.5E-10 97 H4xH9119P2 8.2E-11* 96 H4H9057P2 NBI/Enhancer -57
H4H9068P2 NBI/Enhancer -43 H4xH9037P 7.8E-11* 95 H2aM7780N 9.1E-11*
100 H2aM7788N 6.5E-11* 100 H1M7789N NBI 9 H2aM7790N 1.9E-10 99
H2aM7791N NBI/Enhancer -45 H2aM7794N 2.3E-10 99 H2aM7795N2 6.9E-11*
99 H2aM7796N 1.3E-09 60 H2aM7798N 7.3E-11* 100 H1M7799N 5.9E-11*
100 H1M7800N 6.5E-11* 99 H4xH8992P 1.6E-10 97 H4xH8999P 1.8E-10 92
H4xH9008P 1.3E-09 93 Isotype control 1 NBI 19 Isotype control 2 NBI
35 Isotype control 2 NBI -18 Isotype control 2 NBI -11 Assay
theoretical bottom: for blocking ELISA with biot-hPD-1-mFc constant
and hPD-L1 coat is 1.0E-10M *Below theoretical bottom of the assay;
NT - not tested; NBI - non-blocker; NBI/Enhancer -
non-blocker/enhancer; IC - inconclusive
TABLE-US-00014 TABLE 14 ELISA blocking of human PD-1 binding to
human PD-L2 by anti-PD-1 antibodies Blocking 100 pM of 100 nM
Antibody blocking hPD-1-mFc binding 100 pM hPD-1-mFc to hPD-L2-hFc,
binding to Antibody IC.sub.50 (M) hPD-L2-hFc, % blocking H4xH9048P2
1.4E-10 98 H2aM7795N2 2.6E-10 100 H2aM7798N 1.3E-10 100 H4xH9008P
1.3E-09 94 Isotype control 2 NBI -27 Assay theoretical bottom:
blocking ELISA with hPD-1-mFc constant and hPD-L2 coat is 5.0E-11M
NBI--non-blocker
[0208] As indicated in Table 12, in the first assay format, 23 of
the 27 anti-PD-1 antibodies blocked 1.5 nM of hPD-1-mFc from
binding to hPD-L1-hFc with IC.sub.50 values ranging from 190 pM to
3.3 nM with the percent maximum blockage ranging from 67% to 100%.
One antibody, H2aM7796N, was identified as a non-blocker. Three
anti-PD-1 antibodies (H4H9068P2, H1M7789N, and H2aM7791N) were
identified as non-blockers/enhancers.
[0209] As shown in Table 13, in the second assay format, 23 of the
27 anti-PD-1 antibodies blocked 200 pM of biot-hPD-1-hFc from
binding to hPD-L1-mFc with IC.sub.50 values ranging from 59 pM to
1.3 nM with maximum percent blockade ranging from 60% to 101%. One
antibody, H1M7789N, was identified as a non-blocker. Three
anti-PD-1 antibodies (H4H9057P2, H4H9068P2, and H2aM7791N) were
identified as non-blockers/enhancers.
[0210] In the third assay format as shown in Table 14, four
anti-PD-1 antibodies of the invention, and an Isotype control were
tested. All 4 anti-PD-1 antibodies of the invention blocked 100 pM
(fixed concentration) of hPD-1-mFc from binding to plate-coated
hPD-L2-hFc with IC.sub.50 values ranging from 0.13 nM to 1.3 nM and
with maximum percent blockade ranging from 94% to 100%.
Example 5
Blocking of PD-1 Binding to PD-L1 as Determined by Biosensor Assay
and by Surface Plasmon Resonance
[0211] Inhibition of human PD-1 from binding to human PD-L1 by
different anti-PD-1 monoclonal antibodies was studied either using
real time bio-layer interferometry assay on an Octet Red96
biosensor instrument (Fortebio Inc.) or using a real-time surface
plasmon resonance biosensor assay on a Biacore 3000 instrument.
[0212] Inhibition studies for anti-PD-1 monoclonal antibodies
expressed with a mouse Fc were performed on an Octet Red 96
instrument. First, 100 nM of a recombinant human PD-1 expressed
with a C-terminal mouse IgG2a Fc tag (hPD-1-mFc; SEQ ID NO: 323)
was incubated with 500 nM of each anti-PD-1 monoclonal antibody for
at least 1 hour before running the inhibition assay. Around 0.8 nm
to 1.2 nm of recombinant human PD-L1 expressed with a C-terminal
human IgG1 Fc tag (hPD-L1-hFc; SEQ ID NO: 325) was captured using
anti-human IgG Fc capture Octet biosensor. The Octet biosensors
coated with hPD-L1-hFc were then dipped into wells containing the
mixture of hPD-1-mFc and different anti-PD-1 monoclonal antibodies.
The entire experiment was performed at 25.degree. C. in Octet HBST
buffer (0.01 M HEPES pH7.4, 0.15M NaCl, 3 mM EDTA, 0.05% v/v
Surfactant P20, 0.1 mg/mL BSA) with the plate shaking at a speed of
1000 rpm. The biosensors were washed in Octet HBST buffer in
between each step of the experiment. The real-time binding
responses were monitored during the entire course of the experiment
and the binding response at the end of every step was recorded.
Binding of hPD-1-mFc to the captured hPD-L1-hFc was compared in the
presence and absence of different anti-PD-1 monoclonal antibodies
and was used to determine the blocking behavior of the tested
antibodies as shown in Table 15.
TABLE-US-00015 TABLE 15 Inhibition of human PD-L1 binding to PD-1
by anti-PD-1 monoclonal antibodies expressed with mouse Fc as
measured on an Octet Red 96 instrument Amount of Binding of the
mixture of hPD-L1-hFc 100 nM hPD-1-mFc and Anti-PD-1 Captured 500
nM anti-PD-1 mono- % antibody (nm) clonal antibody (nm) Blocking No
Antibody 0.77 0.07 0 H2aM7780N 1.07 -0.01 114 H2aM7788N 0.74 0.00
100 H1M7789N 0.80 0.05 29 H2aM7790N 0.90 -0.01 114 H2aM7791N 1.17
0.23 -229 H2aM7794N 0.87 -0.01 114 H2aM7795N 0.28 -0.01 114
H2aM7796N 0.82 -0.02 129 H2aM7798N 0.85 0.01 86 H1M7799N 0.79 0.00
100 H1M7800N 0.96 0.00 100
[0213] As shown in Table 15, 9 of the 11 anti-PD-1 antibodies
tested on the Octet Red 96 instrument demonstrated strong blocking
of hPD-1-mFc from binding to hPD-L1-hFc ranging from 86% to
complete blockade of binding. One anti-PD-1 antibody (H1M7789N)
tested showed weaker blocking of hPD-1-mFc binding to hPD-L1-hFc
with 29% blockade. One antibody (H2aM7791N) tested demonstrated the
ability to enhance the binding of hPD-1-mFc to hPD-L1-hFc.
[0214] Next, inhibition studies for anti-PD-1 monoclonal antibodies
expressed with human Fc were performed on a Biacore 3000
instrument. First, 100 nM of a recombinant human PD-1 expressed
with a C-terminal human IgG1 Fc tag (hPD-1-hFc; SEQ ID: 324) was
incubated with 500 nM of each anti-PD-1 monoclonal antibody for at
least 2 hours before running the inhibition assay. A CM5 Biacore
sensor surface was first derivatized with polyclonal rabbit
anti-mouse antibody (GE Catalog#BR-1008-38) using standard EDC-NHS
chemistry. Around 730 RUs of recombinant human PD-L1 expressed with
a C-terminal mouse IgG2a Fc tag (hPD-L1.mFc; SEQ ID: 326) was then
captured followed by the injection of 100 nM of hPD-1.hFc in the
presence and absence of different anti-PD-1 monoclonal antibodies
at a flow rate of 25 .mu.L/min for 3 minutes. The entire experiment
was performed at 25.degree. C. in running buffer comprised of 0.01
M HEPES pH7.4, 0.15M NaCl, 3mM EDTA, 0.05% v/v Surfactant Tween-20
(HBS-ET running buffer).The real-time binding responses were
monitored during the entire course of the experiment and the
binding response at the end of every step was recorded. Binding of
hPD-1-hFc to the captured hPD-L1-mFc was compared in the presence
and absence of different anti-PD-1 monoclonal antibodies and was
used to determine the blocking behavior of the tested antibodies as
shown in Table 16.
TABLE-US-00016 TABLE 16 Inhibition of human PD-L1 binding to PD-1
by anti-PD-1 monoclonal antibodies expressed with human Fc as
measured on a Biacore 3000 instrument Binding of the mixture of 100
nM hPD-1.hFc Anti-PD-1 500 nM of anti- and 500 nM anti- monoclonal
PD-1 monoclonal PD-1 monoclonal % antibody antibody (RU) antibody
(RU) Blocking No Antibody N/A 100 .+-. 1.78 N/A H4H9019P -2 -1 101
H4xH9034P -4 -5 105 H4xH9035P -3 -4 104 H4xH9037P -4 -4 104
H4xH9045P -4 -5 105 H4H9048P2 -7 9 91 H4H9057P2 58 57 43 H4H9068P2
-2 365 -265 H4xH9119P2 -5 -5 105 H4xH9120P2 1 0 100 H4xH9128P2 -5
-5 105 H4xH9135P2 -3 -3 102 H4xH9145P2 -8 -6 106 H4xH8992P 3 2 98
H4xH8999P 1 0 100 H4xH9008P 0 1 99 H4H7795N2 -5 -6 106 H4H7798N -6
-6 106 H4H9008P -7 -7 107 H4H9048P2 -4 6 94
[0215] As shown in Table 16, 18 out of 20 anti-PD-1 antibodies of
the invention tested on the Biacore 3000 instrument demonstrated
strong blocking of hPD-1-hFc from binding to hPD-L1-mFc with the
blockade ranging from 96% to 100%. One antibody demonstrated the
ability to enhance the binding of hPD-1-hFc binding to hPD-L1-mFc.
In this study, one of the tested antibodies of the invention
(H4H9057P2) demonstrated non-specific background binding to the
anti-mouse Fc capture surface.
Example 6
Octet Cross-Competition Between Anti-PD-1 Antibodies
[0216] Binding competition between anti-PD-1 monoclonal antibodies
was determined using a real time, label-free bio-layer
interferometry assay on an Octet RED384 biosensor (Pall ForteBio
Corp.). The entire experiment was performed at 25.degree. C. in
0.01 M HEPES pH7.4, 0.15M NaCl, 3 mM EDTA, 0.05% v/v Surfactant
Tween-20, 0.1 mg/mL BSA (Octet HBS-ET buffer) with the plate
shaking at the speed of 1000 rpm. To assess whether 2 antibodies
were able to compete with one another for binding to their
respective epitopes on a recombinantly expressed human PD-1 with a
C-terminal myc-myc-hexahistidine tag (hPD-1-MMH; SEQ ID: 321),
around 0.1 nM of hPD-1-MMH was first captured onto anti-Penta-His
antibody coated Octet biosensor tips (Pall ForteBio Corp.,
#18-5079) by submerging the tips for 5 minutes into wells
containing a 50 .mu.g/mL solution of hPD-1-MMH. The antigen
captured biosensor tips were then saturated with the first
anti-PD-1 monoclonal antibody (subsequently referred to as mAb-1)
by dipping into wells containing 50 .mu.g/mL solution of mAb-1 for
5 minutes. The biosensor tips were then subsequently dipped into
wells containing a 50 .mu.g/mL solution of a second anti-PD-1
monoclonal antibody (subsequently referred to as mAb-2). The
biosensor tips were washed in Octet HBS-ET buffer in between every
step of the experiment. The real-time binding response was
monitored during the course of the experiment and the binding
response at the end of every step was recorded. The response of
mAb-2 binding to hPD-1-MMH pre-complexed with mAb-1 was compared
and competitive/non-competitive behavior of different anti-PD-1
monoclonal antibodies was determined. Results are summarized in
Table 17 (*Self-competing mAb2s are not listed).
TABLE-US-00017 TABLE 17 Cross-competition between pairs of selected
anti-PD-1 antibodies First antibody applied ("mAb1") mAb2
Antibodies shown to compete with mAb1* H4xH8992P H4xH8999P,
H1M7799N, H2aM7780N, H1M7800N, H2aM7788N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P,
H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P,
H4xH9045P, H2aM7795N H4xH8999P H4xH8992P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7788Nf H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2,
H4xH9120P2, H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2,
H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N,
H4xH9008P H1M7799N H4xH8992P, H4xH8999P, H2aM7780N, H1M7800N,
H2aM7788N, H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2,
H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N,
H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N H2aM7780N H4xH8992P,
H4xH8999P, H1M7799N, H1M7800N, H2aM7788N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P,
H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P,
H4xH9045P, H2aM7795N, H4xH9008P H1M7800N H4xH8992P, H4xH8999P,
H1M7799N, H2aM7780N, H2aM7788N, H2aM7794N, H2aM7798N, H4xH9145P2,
H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P, H4xH9119P2,
H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P, H4xH9045P,
H2aM7795N, H4xH9008P H2aM7788N H4xH8992P, H4xH8999P, H1M7799N,
H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2,
H4xH9120P2, H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2,
H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N,
H2aM7791N H2aM7794N H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7788N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2,
H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N,
H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N, H4xH9008P H2aM7798N
H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N, H1M7800N, H2aM7788N,
H2aM7794N, H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P,
H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P,
H4xH9045P, H2aM7795N, H4xH9008P H4xH9145P2 H4xH8992P, H4xH8999P,
H1M7799N, H2aM7780N, H1M7800N, H2aM7788N, H2aM7794N, H2aM7798N,
H4H9057P2, H4xH9120P2, H4xH9128P2, H4H9019P, H4xH9119P2,
H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P, H4xH9045P,
H4xH9008P H4H9057P2 H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7788N, H2aM7794N, H2aM7798N, H4xH9145P2, H4xH9120P2,
H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N,
H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N, H2aM7791N, H4xH9048P2
H4xH9120P2 H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N, H1M7800N,
H2aM7788N, H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9128P2,
H4H9019P, H4xH9119P2, H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P,
H4xH9037P, H4xH9045P, H4xH9048P2 H4xH9128P2 H4xH8992P, H4xH8999P,
H1M7799N, H2aM7780N, H1M7800N, H2aM7788N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4H9019P, H4xH9119P2,
H4xH9135P2, H4xH9034P, H2aM7790N, H4xH9035P, H4xH9037P, H4xH9045P,
H4xH9008P, H4H9066P2, H4xH9048P2 H4H9019P H4xH8992P, H4xH8999P,
H1M7799N, H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N, H4xH9145P2,
H4H9057P2, H4xH9120P2, H4xH9128P2, H2aM7788N, H4xH9119P2,
H4xH9135P2, H4xH9034P, H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N,
H2aM7791N H4xH9119P2 H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2,
H4xH9128P2, H2aM7788N, H4H9019P, H4xH9135P2, H4xH9034P, H4xH9035P,
H4xH9037P, H4xH9045P, H2aM7795N, H2aM7791N H4xH9135P2 H4xH8992P,
H4xH8999P, H1M7799N, H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H2aM7788N, H4H9019P,
H4xH9119P2, H4xH9034P, H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N,
H2aM7791N H4xH9034P H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2,
H4xH9128P2, H4H9019P, H4xH9119P2, H4xH9135P2, H2aM7788N, H2aM7790N,
H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N, H2aM7791N H2aM7790N
H4xH8992P, H1M7799N, H2aM7780N, H1M7800N, H2aM7788N, H2aM7794N,
H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2,
H4xH9034P, H4xH8999P, H4xH9008P H4xH9035P H4xH8992P, H4xH8999P,
H1M7799N, H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N, H4xH9145P2,
H4H9057P2, H4xH9120P2, H4xH9128P2, H2aM7788N, H4H9019P, H4XH9119P2,
H4xH9034P, H4xH9135P2, H4xH9037P, H4xH9045P, H2aM7795N, H2aM7791N
H4xH9037P H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N, H1M7800N,
H2aM7794N, H2aM7798N, H4xH9145P2, H4H9057P2, H4xH9120P2,
H4xH9128P2, H2aM7788N, H4H9019P, H4xH9119P2, H4xH9034P, H4xH9135P2,
H4xH9035P, H4xH9045P, H2aM7795N, H2aM7791N H4xH9045P H4xH8992P,
H4xH8999P, H1M7799N, H2aM7780N, H1M7800N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4H9057P2, H4xH9120P2, H4xH9128P2, H2aM7788N, H4H9019P,
H4xH9119P2, H4xH9034P, H4xH9135P2, H4xH9035P, H4xH9037P, H2aM7795N,
H2aM7791N H2aM7795N H4xH8992P, H4xH8999P, H1M7799N, H2aM7780N,
H1M7800N, H2aM7794N, H2aM7798N, H4H9057P2, H2aM7788N, H4H9019P,
H4xH9119P2, H4xH9034P, H4xH9135P2, H4xH9035P, H4xH9037P, H4xH9045P,
H2aM7791N H4xH9008P H4xH8999P, H2aM7780N, H2aM7794N, H2aM7798N,
H4xH9145P2, H4xH9128P2, H2aM7790N, H4H9068P2, H1M7799N, H4xH9048P2
H2aM7791N H2aM7788N, H4H9057P2, H4H9019P, H4xH9119P2, H4xH9135P2,
H4xH9034P, H4xH9035P, H4xH9037P, H4xH9045P, H2aM7795N H4H9068P2
H4xH9128P2, H4xH9008P, H1M7789N, H4xH9048P2 H1M7789N H4xH9008P,
H4H9068P2, H4xH9048P2 H4xH9048P2 H4H9057P2, H4xH9120P2, H4xH9128P2,
H4H9019P, H4xH9008P, H4H9068P2, H1M7799N
[0217] A second binding competition between a panel of selected
anti-PD-1 monoclonal antibodies was determined using a real time,
label-free bio-layer interferometry assay on an Octet HTX biosensor
(Pall ForteBio Corp.). The entire experiment was performed at
25.degree. C. in 0.01 M HEPES pH7.4, 0.15M NaCl, 3 mM EDTA, 0.05%
v/v Surfactant Tween-20, 0.1 mg/mL BSA (Octet HBS-ET buffer) with
the plate shaking at the speed of 1000 rpm. To assess whether 2
antibodies were able to compete with one another for binding to
their respective epitopes on the hPD-1-MMH, around 0.25 nm of
hPD-1-MMH was first captured onto anti-Penta-His antibody coated
Octet biosensor tips (Fortebio Inc, #18-5079) by submerging the
tips for 150 seconds into wells containing a 10 .mu.g/mL solution
of hPD-1-MMH. The antigen-captured biosensor tips were then
saturated with a first anti-PD-1 monoclonal antibody (subsequently
referred to as mAb-1) by dipping into wells containing 100 .mu.g/mL
solution of mAb-1 for 5 minutes. The biosensor tips were then
subsequently dipped into wells containing a 100 .mu.g/mL solution
of second anti-PD-1 monoclonal antibody (subsequently referred to
as mAb-2) for 4 minutes. All the biosensors were washed in Octet
HBS-ET buffer in between every step of the experiment. The
real-time binding response was monitored during the course of the
experiment and the binding response at the end of every step was
recorded as shown in FIG. 2. The response of mAb-2 binding to
hPD-1-MMH pre-complexed with mAb-1 was compared and
competitive/non-competitive behavior of different anti-PD-1
monoclonal antibodies was determined. Results are summarized in
Table 18 (*Self-competing mAb2s are not listed).
TABLE-US-00018 TABLE 18 Cross-competition between pairs of selected
anti-PD-1 antibodies First Antibody mAb2 Antibodies Shown applied
("mAb1") to Compete with mAb1* H4H7795N2 H4H7798N H4H7798N
H4H7795N2; H4H9008P H4H9008P H4H7798N; H4H9068P2 H4H9068P2
H4H9008P; H4H9048P2 H4H9048P2 H4H9068P2
[0218] Under the experimental conditions disclosed in this Example,
H4H7795N2 cross-competed with H4H7798N; H4H7798N cross-competed
with H4H7795N2 and H4H9008P; H4H9008P cross-competed with H4H7798N
and H4H9068P2; H4H9068P2 cross-competed with H4H9008P and
H4H9048P2.
Example 7
Antibody Binding to Cells Overexpressing PD-1
[0219] The binding of anti-PD-1 antibodies to a human embryonic
kidney cell line (HEK293; ATCC, #CRL-1573) stably transfected with
full length human PD-1 (amino acids 1 to 289 of accession number
NP_005009.2) (HEK293/hPD-1) was determined by FACS.
[0220] For the assay, adherent cells were detached using trypsin or
enzyme-free dissociation buffer and blocked with complete medium.
Cells were centrifuged and resuspended at a concentration of
2.5-6.times.10 6 cells/mL in cold PBS containing 2% FBS. HEK293
parental and HEK293/hPD-1 cells were then incubated for 15-30min on
ice with 100 nM of each anti-PD-1 antibody. Unbound antibodies were
removed by washing with D-PBS containing 2% FBS, and cells were
subsequently incubated with an allophycocyanin-conjugated secondary
F(ab')2 recognizing either human Fc (Jackson ImmunoResearch,
#109-136-170) or mouse Fc (Jackson ImmunoResearch, #115-136-146)
for 15-30 minutes on ice. Cells were washed with D-PBS containing
2% FBS to remove unbound secondary F(ab')2 and fluorescence
measurements were acquired using either a HyperCyte (IntelliCyt,
Inc.) flow cytometer or an Accuri flow cytometer (BD Biosciences).
Data was analyzed using FlowJo software (Tree Star).
TABLE-US-00019 TABLE 19 FACS binding of anti-PD-1 antibodies to
HEK293/hPD-1 cells and parental HEK293 cells FACS on Ratio of
HEK293 FACS on HEK293/hPD-1 parental HEK293/hPD- to HEK293 Antibody
cells [MFI] 1 cells [MFI] parental cells H1M7789N 262 24166 92.3
H1M7799N 255 6855 26.9 H1M7800N 275 6812 24.7 H2aM7780N 320 23656
73.8 H2aM7788N 305 23112 75.7 H2aM7790N 270 47310 175.5 H2aM7791N
274 4948 18.0 H2aM7794N 270 19127 71.0 H2aM7795N 288 817 2.8
H2aM7796N 297 49755 167.8 H2aM7798N 300 23443 78.1 H4H9019P 111
8610 77.2 H4H9057P2 141 6501 46.1 H4H9068P2 285 1940 6.8 H4xH8992P
358 17502 48.9 H4xH8999P 809 28875 35.7 H4xH9008P 509 26233 51.5
H4xH9034P 147 10115 69.0 H4xH9035P 108 9915 91.7 H4xH9037P 108 8787
81.4 H4xH9045P 95 8884 93.7 H4xH9048P2 102 7196 70.8 H4xH9119P2 109
9142 84.0 H4xH9120P2 109 9975 91.9 H4xH9128P2 135 9081 67.5
H4xH9135P2 114 9380 82.2 H4xH9145P2 226 11552 51.2
[0221] As shown in Table 19, 25 of the 27 anti-PD-1 antibodies of
the invention showed strong binding to the HEK293/hPD-1 cells
compared to binding on the parental HEK293 line. Two antibodies of
the invention (H2aM7795N and H4H9068P2) bound weaker to human PD-1
expressing cells compared to the other antibodies tested.
[0222] To further characterize anti-PD1 antibodies of the
invention, dose-dependent binding to a human embryonic kidney cell
line (HEK293; ATCC, #CRL-1573) stably transfected with full length
human PD-1 (amino acids 1 to 289 of accession number NP_005009.2)
(HEK293/hPD-1) was determined by FACS.
[0223] For the assay, adherent cells were detached using trypsin
and blocked with complete medium. Cells were centrifuged and
resuspended at a concentration of 6.times.10 6 cells/mL in staining
buffer (1% FBS in PBS). To determine the EC.sub.50 and E.sub.max of
the anti-PD1 antibodies, 90 uL of cell suspension was incubated for
30 minutes on ice with a serial dilution of anti-PD-1 antibodies
and controls diluted to a final concentration ranging from 5 pM to
100 nM (no mAb sample was included as negative control) in staining
buffer. Cells were then centrifuged and pellets were washed once
with staining buffer to remove unbound antibodies. Cells were
subsequently incubated for 30 minutes on ice either with an
allophycocyanin-conjugated secondary F(ab')2 recognizing human Fc
(Jackson ImmunoResearch, #109-136-170) or mouse Fc (Jackson
ImmunoResearch, #115-136-071). Cells were centrifuged and pellets
were washed once with staining buffer to remove unbound secondary
F(ab')2 and then fixed overnight with a 1:1 dilution of Cytofix (BD
Biosciences, #554655) and staining buffer. The following day, cells
were centrifuged and pellets were washed once with staining buffer,
resuspended and filtered. Fluorescence measurements were acquired
on Hypercyt.RTM. cytometer and analyzed in ForeCyt.TM. (IntelliCyt;
Albuquerque, N. Mex.) to determine the mean fluorescence
intensities (MFI). The EC.sub.50 values were calculated from a
four-parameter logistic equation over an 11-point response curve
using GraphPad Prism. E.sub.max for each antibody was defined as
the binding at the highest antibody dose (100 nM) tested.
TABLE-US-00020 TABLE 20 Dose dependent FACS binding of anti-PD-1
antibodies to HEK293/hPD-1 cells Max Geom. Mean Antibody EC.sub.50
[M] [MFI] @ 100 nM H2aM7779N 2.59E-09 16832 H2aM7780N 1.69E-09
18415 H2aM7781N 5.67E-10 13740 H2aM7782N 1.26E-09 17302 H2aM7787N
2.40E-09 15744 H2aM7788N 3.21E-10 14827 H2aM7790N 1.71E-10 19196
H2aM7791N No EC.sub.50 determined 1397 H2aM7794N 1.37E-09 16406
H2aM7795N No EC.sub.50 determined 624 H2aM7798N 6.985E-11 20900
H1M7799N 3.318E-11 24405 H1M7800N 4.80E-11 20763 H4xH8992P 5.45E-11
11368 H4xH8999P 5.27E-11 28341 H4H9019P 1.40E-09 29201 H4xH9034P
2.09E-10 32388 H4xH9035P 1.15E-10 28708 H4xH9037P 6.74E-10 36441
H4xH9045P 9.17E-11 24662 H4xH9048P2 6.68E-10 33687 H4H9057P2
2.363E-10 19953 H4H9068P2 No EC.sub.50 determined 639 H4xH9119P2
3.476E-10 37789 H4xH9120P2 4.797E-10 34057 H4xH9128P2 1.551E-09
37167 H4xH9135P2 1.048E-10 32793 H4xH9145P2 2.321E-10 30613 mlgG1
isotype N/A 200 mlgG2a isotype N/A 239 hlgG4 isotype N/A 459
TABLE-US-00021 TABLE 21 Dose dependent FACS binding of anti-PD-1
antibodies to HEK293/hPD-1 cells Max Geom. Mean Antibody EC.sub.50
[M] [MFI] @ 100 nM H4H7795N2 Inconclusive 15188 H4H7798N 5.09E-10
20305 H4H9008P Inconclusive 32230 H4H9048P2 1.60E-09 39774 H1M7789N
Inconclusive 35574 H2aM7796N 4.81E-09 14111 mlgG1 isotype N/A 858
mlgG2a isotype N/A 352 hlgG4 isotype N/A 809
[0224] As shown in Table 20, 25 of 28 anti-PD1 antibodies of the
invention showed dose dependent binding to HEK293/hPD-1 cells with
EC.sub.50 values ranging from 33.18 pM to 2.59 nM and E.sub.max
values ranging from 37,789 to 11,368 MFI. Three anti-PD1 antibodies
of the invention did not demonstrate strong binding to HEK293/hPD-1
cells and therefore an EC.sub.50 value could not be determined.
None of the isotype controls demonstrated any measurable binding in
this assay.
[0225] As shown in Table 21, 3 of 6 anti-PD1 antibodies of the
invention showed dose dependent binding to HEK293/hPD-1 cells with
EC.sub.50 values ranging from 509 pM to 4.81 nM and E.sub.max
values ranging from 39,774 to 14,111 MFI. Three antibodies of the
invention tested bound to HEK293/hPD-1 cells, but did not reach a
plateau. Therefore their precise EC.sub.50 values could not be
determined and their EC.sub.50 values are referred to as
inconclusive. None of the isotype controls demonstrated any
measurable binding in this assay.
Example 8
Blocking of PD-1-Induced T-Cell Down-Regulation in a T-Cell/APC
Luciferase Reporter Assay
[0226] T-cell activation is achieved by stimulating T-cell
receptors (TcR) that recognize specific peptides presented by major
histocompatibility complex class I or II proteins on
antigen-presenting cells (APC). Activated TcRs in turn initiate a
cascade of signaling events that can be monitored by reporter genes
driven by transcription factors such as activator-protein 1 (AP-1),
Nuclear Factor of Activated T-cells (NFAT) or Nuclear factor
kappa-light-chain-enhancer of activated B cells (NFKb). T-cell
response is modulated via engagement of co-receptors expressed
either constitutively or inducibly on T-cells. One such receptor is
PD-1, a negative regulator of T-cell activity. PD-1 interacts with
its ligand, PD-L1, which is expressed on target cells including
APCs or cancer cells, and acts to deliver inhibitory signals by
recruiting phosphatases to the TcR signalosome, resulting in the
suppression of positive signaling.
[0227] The ability of anti-PD-1 antibodies to antagonize
PD-1/PD-L1-mediated signaling through the PD-1 receptor in human T
cell lines was assessed using an in vitro cell based assay shown in
FIG. 1. The bioassay was developed to measure T cell signaling
induced by interaction between APC and T cells by utilizing a mixed
culture derived from two mammalian cell lines: Jurkat cells (an
immortalized T cell line) and Raji cells (a B cell line). For the
first component of the bioassay, Jurkat Clone E6-1 cells (ATCC,
#TIB-152) were transduced with the Cignal Lenti AP-1 Luc Reporter
(Qiagen--Sabiosciences, #CLS-011L) as per the manufacturer's
instructions. The lentivirus encodes the firefly luciferase gene
under the control of a minimal CMV promoter, tandem repeats of the
TPA-inducible transcriptional response element (TRE) and a
puromycin resistance gene. The engineered Jurkat cell line was
subsequently transduced with a PD-1 chimera comprising the
extracellular domain of human PD-1 (amino acids from 1 to 170 of
human PD1; accession number NP_005009.2) and the trans-membrane and
cytoplasmic domains of human CD300a (amino acids from 181 to 299 of
human CD300a; accession number NP_009192.2). The resulting stable
cell line (Jurkat/AP1-Luc/hPD1-hCD300a) was selected and maintained
in RPMI/10% FBS/penicillin/streptomycin/glutamine supplemented with
500 ug/mL G418+1 ug/mL puromycin.
[0228] For the second component of the bioassay, Raji cells (ATCC,
#CCL-86) were transduced with human PD-L1 gene (amino acids 1-290
of accession number NP_054862.1) that had been cloned into a
lentiviral (pLEX) vector system (Thermo Scientific Biosystems,
#OHS4735). Raji cells, positive for PD-L1 (Raji/hPD-L1) were
isolated by FACS using a PD-L1 antibody and maintained in
Iscove/10% FBS/penicillin/streptomycin/glutamine supplemented with
1 ug/mL puromycin.
[0229] To simulate the APC/T cell interaction, a bispecific
antibody composed of one Fab arm that binds to CD3 on T cells and
the other one Fab arm binding that binds to CD20 on Raji cells
(CD3xCD20 bispecific antibody; e.g., as disclosed in US20140088295)
was utilized. The presence of the bispecific molecule in the assay
results in the activation of the T cell and APC by bridging the CD3
subunits on T-cells to CD20 endogenously expressed on Raji cells.
Ligation of CD3 with anti-CD3 antibodies has been demonstrated to
lead to activation of T cells. In this bioassay, antibodies
blocking the PD1/PD-L1 interaction rescue T-cell activity by
disabling the inhibitory signaling and subsequently leading to
increased AP1-Luc activation.
[0230] In the luciferase-based bioassay, RPMI1640 supplemented with
10% FBS and penicillin/streptomycin/glutamine was used as assay
medium to prepare cell suspensions and antibody dilutions to carry
out the screening of anti-PD1 monoclonal antibodies (mAbs). On the
day of the screening, EC50 values of anti-PD1 mAbs, in the presence
of a fixed concentration of CD3xCD20 bispecific antibody (30 pM),
as well as the EC50 of the bispecific antibody alone, were
determined. In the following order, cells and reagents were added
to 96 well white, flat-bottom plates. For the anti-PD1 mAb EC50
determinations, first a fixed concentration of CD3xCD20 bispecific
antibody (final 30 pM) was prepared and added to the microtiter
plate wells. Then 12-point serial dilutions of anti-PD1 mAbs and
controls were added (final concentrations ranging from 1.7 pM to
100 nM; plus wells with assay medium alone). For the bispecific
antibody (alone) EC.sub.50 determination, the bispecific antibody,
at final concentrations ranging from 0.17 pM to 10 nM (plus wells
with assay medium alone), was added to the microtiter plate wells.
Subsequently, a 2.5.times.10 6/mL Raji/hPD-L1 cell suspension was
prepared and 20 uL per well was added (final cell number/well
5.times.10 4 cells). Plates were left at room temperature (15-20
minutes), while a suspension of 2.5.times.10 6/mL of
Jurkat/AP1-Luc/hPD1(ecto)-hCD300a(TM-Cyto) was prepared. 20 uL of
the Jurkat suspension (final cell number/well 5.times.10 4 cells)
was added per well. Plates containing the co-culture were incubated
for 5 to 6 hours at 37.degree. C./5% CO.sub.2. Samples were tested
in duplicates and luciferase activity was then detected after the
addition of ONE-Glo.TM. (Promega, #E6051) reagent and relative
light units (RLUs) were measured on a Victor luminometer.
[0231] RLU values for each screened antibody were normalized by
setting the assay condition with fixed (30 pM) concentration of the
CD3/CD20 bispecific antibody, but without anti-PD-1 antibody to
100%. This condition corresponds to the maximal AP1-Luc response
elicited by the bispecific molecule in the presence of the
PD-1/PD-L1 inhibitory signal. Upon addition of the anti-PD-1
antibody, the inhibitory signal is suppressed, and the increased
stimulation is shown here as E.sub.max, the percentage increase in
the signal in the presence of the highest antibody dose tested (100
nM). To compare potency of the anti-PD1 antibodies tested, the
concentration of antibody at which the normalized RLU value reached
150% activation was determined from a four-parameter logistic
equation over a 12-point response curve using GraphPad Prism. The
results are summarized in Table 22 and Table 23, respectively.
TABLE-US-00022 TABLE 22 Anti-PD1 antibody blocking PD-1/ PD-L1
dependent inhibition of AP1-Luc signaling in Experiment 1
Antagonistic assay Antagonistic assay Concentration (M) of
E.sub.max mean Antibody at 150% [%] @ 100 nM Antibody activation
Experiment 1 Experiment 1 H1M7789N N/A 135 H1M7799N 2.97E-08 183
H1M7800N 1.65E-08 182 H2aM7779N 8.92E-09 214 H2aM7780N 6.52E-09 228
H2aM7781N 6.70E-09 230 H2aM7782N 9.96E-09 215 H2aM7787N 1.38E-08
215 H2aM7788N 4.72E-09 189 H2aM7790N 5.24E-09 234 H2aM7791N N/A 103
H2aM7794N 4.09E-08 170 H2aM7795N N/A 109 H2aM7796N N/A 121
H2aM7798N 7.99E-10 239 H4H9019P 1.79E-08 180 H4xH9034P 2.62E-09 202
H4xH9035P 1.20E-09 227 H4xH9037P 2.82E-09 195 H4xH9045P 2.23E-08
176 H4xH9048P2 N/A 138 H4H9057P2 2.68E-08 212 H4H9068P2 N/A 102
H4xH9119P2 1.11E-08 163 H4xH9120P2 1.10E-08 166 H4xH9128P2 3.99E-09
187 H4xH9135P2 1.55E-09 193 H4xH9145P2 2.40E-09 185 H4xH8992P
5.32E-09 178 H4xH8999P 8.63E-10 217 H4H7798N 1.54E-09 202 mlgG1
isotype control N/A 92 mlgG2a isotype control N/A 91 hlgG4 isotype
control N/A 94 N/A = not applicable because at the concentrations
tested these antibodies did not activate 150%
TABLE-US-00023 TABLE 23 Anti-PD1 antibody blocking PD-1/PD-L1
dependent inhibition of AP1-Luc signaling in Experiment 2
Antagonistic assay Antagonistic Concentration (M) assay E.sub.max
of Antibody at mean [%] 150% activation @ 100 nM Antibody
Experiment 2 Experiment 2 H4H7795N2 N/A 110 H4H7798N 1.59E-10 343
H4H9008P 9.84E-08 150 H4H9048P N/A 134 hlgG4 isotype control N/A 98
N/A= not applicable because at the concentrations tested these
antibodies did not activate 150%
[0232] As shown in Table 22, 25 out of the 31 anti-PD-1 antibodies
of the invention tested blocked PD-1/PD-L1 inhibition with
E.sub.max values ranging from 239 to 163. Six out of the 31
anti-PD-1 antibodies of the invention did not demonstrate
substantial blockade of PD1/PD-L1 interaction when tested in this
assay.
[0233] As shown in Table 23, 2 out of the 4 anti-PD-1 antibodies of
the invention tested blocked PD-1/PD-L1 inhibition with E.sub.max
values of 150 and 343%, respectively. 2 out of the 4 anti-PD-1
antibodies of the invention did not demonstrate substantial
blockade of PD1/PD-L1 interaction when tested in this assay.
Example 9
In Vivo Efficacy of Anti-PD-1 Antibodies
[0234] To determine the effect of a select number of anti-PD-1
antibodies of the invention in a relevant in vivo model, three
MC38.ova tumor growth studies, involving subcutaneous injection of
tumor cells and started on different days, were conducted in mice
that were homozygous for the expression of the extracellular domain
of human PD-1 in place of extracellular domain of mouse PD-1 (PD-1
Humin mice) on a 75% C57/BI6/25% 129 strain background.
[0235] For the studies, mice were divided evenly according to body
weight into 5 treatment or control groups for Study 1 (5 mice per
group), 8 treatment or control groups for Study 2 (5 mice per
group), and 5 treatment or control groups for Study 3 (7 mice per
group). At day 0, mice were anesthetized by isoflurane inhalation
and then injected subcutaneously into the right flank with
5.times.10.sup.5 MC38.ova cells in suspension of 100 uL of DMEM for
Study 1 or 1.times.10.sup.6 MC38.ova cells in suspension of 100 uL
of DMEM for Study 2 and Study 3. For Study 1, treatment groups were
intraperitoneally injected with 200 ug of either one of three
anti-PD-1 antibodies of the invention, or an isotype control
antibody with irrelevant specificity on days 3, 7, 10, 14, and 17
of the experiment, while one group of mice was left untreated. For
Study 2, treatment groups were intraperitoneally injected with
either one of three anti-PD-1 antibodies of the invention at 10
mg/kg or 5 mg/kg per/dose, one antibody of the invention
(H4H7795N2) at 10 mg/kg per dose, or an isotype control antibody
with irrelevant specificity at 10 mg/kg on days 3, 7, 10, 14, and
17 of the experiment. For Study 3, treatment groups were
intraperitoneally injected with either one of two anti-PD-1
antibodies of the invention at 5 mg/kg or 2.5 mg/kg per/dose, or an
isotype control antibody with irrelevant specificity at 5 mg/kg on
days 3, 7, 10, 14, and 17 of the experiment. Experimental dosing
and treatment protocol for groups of mice are shown in Table
24.
TABLE-US-00024 TABLE 24 Experimental dosing and treatment protocol
for groups of mice Dosage amount Study Samples at each dosage #
Tested time point Dosing interval 1 Isotype Control 200 ug Days 3,
7, 10, 14, 17 No treatment N/A N/A H4H7798N 200 ug Days 3, 7, 10,
14, 17 H4H7795N2 200 ug Days 3, 7, 10, 14, 17 H4H9008P 200 ug Days
3, 7, 10, 14, 17 2 Isotype Control 10 mg/kg Days 3, 7, 10, 14, 17
H4H7795N2 10 mg/kg Days 3, 7, 10, 14, 17 H4H7798N 10 mg/kg Days 3,
7, 10, 14, 17 H4H7798N 5 mg/kg Days 3, 7, 10, 14, 17 H4H9048P2 10
mg/kg Days 3, 7, 10, 14, 17 H4H9048P2 5 mg/kg Days 3, 7, 10, 14, 17
H4H9008P 10 mg/kg Days 3, 7, 10, 14, 17 H4H9008P 5 mg/kg Days 3, 7,
10, 14, 17 3 Isotype Control 5 mg/kg Days 3, 7, 10, 14, 17 H4H7798N
5 mg/kg Days 3, 7, 10, 14, 17 H4H7798N 2.5 mg/kg Days 3, 7, 10, 14,
17 H4H9008P 5 mg/kg Days 3, 7, 10, 14, 17 H4H9008P 2.5 mg/kg Days
3, 7, 10, 14, 17
[0236] For the studies, average tumor volumes determined by caliper
measurements and percent survival at Day 14 or 17 and Day 23 or 24
of each experiment for each treatment group were recorded. In
addition, the number of tumor-free mice were also assessed at the
end of the study (Day 42 for Study 1 and Day 31 for Study 2 and
Study 3). Results, expressed as mean tumor volume
(mm.sup.3)(.+-.SD), percent survival, and number of tumor-free mice
are shown in Table 23 for Study 1, Table 3 for Study 2, and Table 4
for Study 3.
TABLE-US-00025 TABLE 25 Mean tumor volume, percent survival and
numbers of tumor free mice in each treatment group from in vivo
tumor Study 1 Tumor Volume, mm.sup.3 Tumor-Free mean (.+-.SD)
Survival, % Mice Day 17 Day 23 Day 17 Day 23 Day 42 Treatment 200
200 200 200 200 group (n = 5) ug/mouse ug/mouse ug/mouse ug/mouse
ug/mouse No treatment 189 (.+-.110) 554 (.+-.317) 100% 100% 1/5
(20%) Isotype control 86 (.+-.114) 515 (.+-.859) 100% 60% 2/5 (40%)
H4H7798N 0 (0) 0 (0) 100% 100% 5/5 (100%) H4H9008P 14 (.+-.19) 205
(.+-.312) 100% 100% 3/5 (60%) H4H7795N2 89 (.+-.176) 445 (.+-.889)
100% 80% 3/5 (60%)
[0237] As shown in Table 25 for Study 1, mice treated with one
antibody of the invention, H4H7798N did not develop any detectable
tumors during the course of the study. Mice treated with H4H9008P
exhibited a sustained reduced tumor volume as compared to controls
at days 17 and 24 of the study with 3 out of 5 mice or 4 out of 5
mice being tumor free by the end of the experiment, respectively.
In contrast, treatment with one of the anti-PD1 antibodies,
H4H7795N2, did not demonstrate significant efficacy in reducing
tumor volume in this study as compared to controls. By day 23 of
the study, 1 out of 5 mice died in the H4H7795N2 group, and 2 out
of 5 mice died in the isotype control treatment group. In
non-treatment group and isotype control group some mice exhibited
spontaneous regression of tumors (1 out of 5 mice and 2 out of 5
mice, respectively).
TABLE-US-00026 TABLE 26 Mean tumor volume, percent survival and
numbers of tumor free mice in each treatment group from in vivo
tumor Study 2 Tumor Volume, mm.sup.3 Tumor-Free Treatment mean
(.+-.SD) Survival, % Mice group Days 17 Day 24 Day 17 Day 24 Day 31
(n = 5) 5 mg/kg 10 mg/kg 5 mg/kg 10 mg/kg 5 mg/kg 10 mg/kg 5 mg/kg
10 mg/kg 5 mg/kg 10 mg/kg Isotype N/A 449 (.+-.434) N/A 824
(.+-.858) N/A 100% N/A 60% N/A 1/5 control (20%) H4H7798N 17
(.+-.38) 0 (0) 104 (.+-.233) 0 (0) 100% 100% 100% 100% 4/5 5/5/
(80%) (100%) H4H9008P 91 (.+-.204) 12 (.+-.28) 228 (.+-.509) 96
(.+-.215) 100% 100% 80% 100% 4/5 4/5 (80%) (80%) H4H9048P2 94
(.+-.160) 10 (.+-.21) 328 (.+-.559) 67 (.+-.150) 100% 100% 80% 100%
3/5 4/5 (60%) (80%) H4H7795N2 N/A 124 (.+-.209) N/A 359 (.+-.657)
N/A 100% N/A 80% N/A 2/5 (40%)
[0238] As shown in Table 26 for Study 2, mice treated with one
antibody of the invention, H4H7798N at 10 mg/kg did not develop
detectable tumors during the course of the study. Groups of mice
treated with 10 mg/kg of either H4H9008P or H4H9048P2 exhibited
substantially reduced tumor volume as compared to controls at days
17 and 24 of the study. Four out of 5 mice in each group treated
with 10 mg/kg of either H4H9008P or H4H9048P2 were tumor free at
Day 31, whereas in the isotype control treatment group only 1 out
of 5 animals was tumor free as a result of spontaneous tumor
regression. One antibody tested at 10 mg/kg, H4H7795N2,
demonstrated substantially reduced tumor volume as compared to
controls at days 17 and 24 of the study, but this antibody was the
least efficacious anti-PD1 antibody with only 2 out of 5 mice
surviving at the end of the experiment.
[0239] A dose-dependent response in tumor suppression at the tested
doses (5 mg/kg and 10 mg/kg) was observed in groups treated with
H4H7798N, H4H9008P, and H4H9048P2. H4H7798N or H4H9008P therapy at
5 mg/kg was less efficacious, with 4 out of 5 tumor-free mice at
the end of experiment on day 21, whereas 5 out of 5 mice remained
tumor-free in both 10 mg/kg dose groups of H4H7798N, and
H4H9008P.
[0240] Dunett's test in 2 way ANOVA multiple comparisons revealed
that the differences in tumor growth between the group treated with
isotype control antibody at 10 mg/kg as reference and the groups
treated at 10 mg/kg with either H4H7798N, H4H9008P, or H4H9048P2
were statistically significant with p value<0.005. The
differences in tumor growth between the group treated with isotype
control antibody at 10 mg/kg as reference and the groups treated at
5 mg/kg with either H4H7798N, H4H9008P, or H4H9048P2 were also
statistically significant with a p value<0.05.
TABLE-US-00027 TABLE 27 Mean tumor volume, percent survival and
numbers of tumor free mice in each treatment group from in vivo
tumor Study 3 Tumor Volume, mm.sup.3 mean (.+-.SD) Survival, %
Tumor-Free Mice Treatment Days 14 Day 21 Day 14 Day 21 Day 31 group
(n = 7) 2.5 mg/kg 5 mg/kg 2.5 mg/kg 5 mg/kg 2.5 mg/kg 5 mg/kg 2.5
mg/kg 5 mg/kg 2.5 mg/kg 5 mg/kg Isotype control N/A 94 (.+-.44) N/A
405 (.+-.326) N/A 100% N/A 86% N/A 0/7 (0%) H4H7798N 0 (0) 0 (0) 19
(.+-.51) 13 (.+-.35) 100% 100% 100% 100% 6/7 6/7 (86%) (86%)
H4H9008P 41 (.+-.68) 7 (.+-.20) 87 (.+-.123) 16 (.+-.42) 100% 100%
100% 100% 4/7 6/7 (57%) (86%)
[0241] As shown in Table 27 for Study 3, 6 out or 7 mice treated
with one antibody of the invention, H4H7798N, or another antibody
of the invention, H4H9008P, at 5 mg/kg were tumor free at the end
of the experiment, whereas there were no tumor free animals in the
isotype control group. One tumor-bearing mouse in the IgG4 control
group died on post-implantation day 17. Only 4 out of 7 mice
treated with H4H9008P at 2.5 mg/kg dose remained tumor free at the
end of the experiment. The difference in tumor volumes at day 21
between anti-PD-1 antibodies tested and an isotype control group
was statistically significant as determined by one-way ANOVA with
Dunnett's multiple comparison post-test with p<0.01. All four
anti-PD-1 antibodies were equally more efficacious at the 5 mg/kg
dose than at the 2.5 mg/kg dose.
Example 10
Anti-Tumor Effects of a Combination of an Anti-PD-1 Antibody and a
VEGF Antagonist in a Mouse Early-Treatment Tumor Model
[0242] An early-treatment tumor model was developed to test the
efficacy of a combination of an anti-PD-1 antibody and a VEGF
antagonist. In this model, the combination therapy is administered
shortly after tumor implantation. The experiment also used an
anti-PD-L1 antibody alone and in combination with the VEGF
antagonist. The anti-PD-1 antibody used in this experiment was
anti-mouse PD-1 clone "RPMI-14" with rat IgG2b (Bio X Cell, West
Lebanon, N.H.). The VEGF antagonist used in this experiment was
aflibercept (a VEGF receptor-based chimeric molecule, also known as
"VEGF-trap" or "VEGFR1R2-Fc.DELTA.C1(a)," a full description of
which is provided elsewhere herein). The anti-PD-L1 antibody used
in this experiment was an anti-PD-L1 monoclonal antibody with
V.sub.H/V.sub.L sequences of antibody "YW243.55S70" according to
US20100203056A1 (Genentech, Inc.), with mouse IgG2a and which was
cross-reactive with mouse PD-L1.
[0243] For this experimental model, 1.0.times.10.sup.6 Colon-26
tumor cells were implanted sub-cutaneously into BALB/c mice at Day
0. Starting on Day 3, prior to the establishment of measurable
tumors, mice were treated with one of the mono- or combination
therapies, or control combination, as set forth in Table 28.
TABLE-US-00028 TABLE 28 Experimental dosing and treatment groups
Treatment Group First Agent Second Agent Control IgG2a isotype
control hFc control Combination (250 .mu.g, IP) (250 .mu.g, SC)
VEGF Trap only IgG2a isotype control Aflibercept (250 .mu.g, IP)
(10 mg/kg, SC) anti-PD-1 only anti-PD-1 mAb RPMI-14 hFc control
(250 .mu.g, IP) (250 .mu.g, SC) anti-PD-L1 only anti-PD-L1 mAb hFc
control (250 .mu.g, IP) (250 .mu.g, SC) VEGF Trap + anti-PD-1 mAb
RPMI-14 Aflibercept anti-PD-1 (250 .mu.g, IP) (10 mg/kg, SC) VEGF
Trap + anti-PD-L1 mAb Aflibercept anti-PD-L1 (250 .mu.g, IP) (10
mg/kg, SC)
[0244] The various therapies were administered at five different
time points over a two week period (i.e., injections at Day 3, Day
6, Day 10, Day 13 and Day 19).
[0245] Animals in each therapy group were evaluated in terms of
tumor incidence, tumor volume, median survival time, and number of
tumor-free animals at Day 50. The extent of tumor growth is
summarized in FIG. 2 (tumor growth curves) and FIG. 3 (tumor volume
at Day 28). Results are also summarized in Table 29.
TABLE-US-00029 TABLE 29 Tumor-free mice in treatment groups No. of
Tumor-Free Treatment Group Animals by Day 50 Control Combination
0/10 VEGF Trap only 3/10 anti-PD-1 only 4/10 anti-PD-L1 only 5/10
VEGF Trap + anti-PD-1 7/10 VEGF Trap + anti-PD-L1 9/10
[0246] Tumor growth was substantially reduced in animals treated
with the combination of VEGF Trap+anti-PD-1 antibody as compared
with treatment regimens involving either therapeutic agent alone
(see FIGS. 2 and 3). Furthermore, survival was substantially
increased in the VEGF Trap+anti-PD-1 antibody group, with 70% of
animals surviving to at least day 50 after tumor implantation. By
contrast, for the anti-PD-1 and VEGF Trap monotherapy groups,
survival to Day 50 was only 40% and 30% respectively (see FIG. 3
and Table 29).
Example 11
Clinical Trial Study of Repeat Dosing with Anti-PD-1 Antibody as
Single Therapy and in Combination with Other Anti-Cancer Therapies
in Patients with Advanced Malignancies
[0247] This is a dose-escalation study of anti-PD-1 antibody, alone
or in combination with radiation therapy, cyclophosphamide, or both
in patients with advanced malignancies. The exemplary anti-PD-1
antibody ("mAb") used in this Example comprises HCVR of SEQ ID NO:
162 and LCVR of SEQ ID NO: 170.
Study Objectives
[0248] The primary objective of the study is to characterize the
safety, tolerability, DLTs of mAb administered IV as monotherapy,
or in combination with targeted radiation (with the intent to have
this serve as an immuno-stimulatory, rather than primarily
tumor-ablative therapy), low-dose cyclophosphamide (a therapy shown
to inhibit regulatory T-cell responses), or both in patients with
advanced malignancies.
[0249] The secondary objectives of the study are: (1) to determine
a recommended phase 2 dose (RP2D) of mAb as monotherapy and in
combination with other anti-cancer therapies (targeted radiation,
low-dose cyclophosphamide, or both); (2) to describe preliminary
antitumor activity of mAb, alone and with each combination partner
(s); (3) to characterize the PK of mAb as monotherapy and in
combination with other anti-cancer therapies (targeted radiation,
low-dose cyclophosphamide, or both); and (4) to assess
immunogenicity of mAb.
Study Design
[0250] Safety will be assessed in separate, standard 3+3 dose
escalation cohorts (in monotherapy, combination with radiation
therapy, combination with cyclophosphamide, and combination with
radiation therapy plus cyclophosphamide). The choice of combination
therapy with radiation, cyclophosphamide, or both will be based on
investigator assessment of the best choice of therapy for an
individual patient in consultation with the sponsor. To be enrolled
in a radiotherapy cohort, a patient must have a lesion that can be
safely irradiated and for which radiation at the limited,
palliative doses contemplated would be considered medically
appropriate, and at least one other lesion suitable for response
evaluation. A patient will be allowed to enroll only if a slot is
available in the cohort for the chosen treatment.
[0251] Patients will undergo screening procedures to determine
eligibility within 28 days prior to the initial administration of
mAb. Following enrollment of patients into a mAb monotherapy
cohort, enrollment of subsequent cohorts will be determined by
occurrence of DLTs in prior cohorts (i.e., no DLT in a cohort of 3
patients, or no more than 1 DLT in an expanded cohort of 6
patients), and the availability of patient slots. The planned
monotherapy dose levels are 1, 3, or 10 mg/kg administered IV every
14 days (2 weeks).
[0252] Once one or both of the 1 mg/kg or 3 mg/kg mAb monotherapy
cohort DLT observation periods are completed without a DLT in a
cohort of 3 patients or with no more than 1 DLT in an expanded
cohort of 6 patients, patients can be enrolled into a cohort
combining cyclophosphamide or radiotherapy with mAb at that
monotherapy dose level. Patients can be enrolled into a combination
mAb+cyclophosphamide/radiotherapy cohort once the DLT observation
periods for both the cohort for that mAb dose
level+cyclophosphamide and the cohort for that mAb dose level+the
same radiotherapy regimen are completed with no DLT in a cohort of
3 patients, or no more than 1 DLT in an expanded cohort of 6
patients.
[0253] Once the 3 mg/kg mAb monotherapy cohort DLT observation
period is completed with no DLT in a cohort of 3 patients, or no
more than 1 DLT in an expanded cohort of 6 patients, a 10 mg/kg mAb
monotherapy cohort may also enroll.
[0254] mAb 3 mg/kg and 10 mg/kg monotherapy cohorts will enroll
only after the requisite number of patients in the prior
monotherapy dose cohort (i.e., 1 mg/kg and 3 mg/kg, respectively)
have cleared the 28 day DLT observation period without a maximum
tolerated dose (MTD) being demonstrated for that dose level. A mAb
1 mg/kg combination treatment cohort will enroll only after
completion of the DLT observation period for the 1 mg/kg
monotherapy cohort. Combination cohorts receiving 3 mg/kg mAb will
enroll only when the requisite number of patients in the respective
1 mg/kg mAb combination cohorts has cleared the DLT observation
period without demonstrating a MTD. Triple combination cohorts
combining mAb with cyclophosphamide and a radiation regimen will
enroll only when the requisite number of patients in both
corresponding double combination cohorts at that dosage level have
cleared the DLT observation period without a MTD being
demonstrated.
[0255] Table 30 summarizes the dose-escalation cohorts in which
patients will be enrolled.
TABLE-US-00030 TABLE 30 Possible Dose-escalation Cohorts n Possible
Assigned Treatment Cohort 3-6 0.3 mg/kg mAb monotherapy (to be
enrolled only if MTD <1 mg/kg mAb) 3-6 1 mg/kg mAb monotherapy
3-6 3 mg/kg mAb monotherapy.sup.a) 3-6 10 mg/kg mAb
monotherapy.sup.b) 3-6 1 mg/kg.sup.a) mAb + radiotherapy (6 Gy
.times. 5) 3-6 1 mg/kg.sup.a) mAb + radiotherapy (9 Gy .times. 3)
3-6 3 mg/kg.sup.b) (or MTD) mAb + cyclophosphamide 3-6 3
mg/kg.sup.b) (or MTD) mAb + radiotherapy (6 Gy .times. 5) 3-6 3
mg/kg.sup.b) (or MTD) mAb + radiotherapy (9 Gy .times. 3) 3-6 3
mg/kg.sup.b) (or MTD) mAb + radiotherapy (6 Gy .times. 5) +
cyclophosphamide 3-6 3 mg/kg.sup.b) (or MTD) mAb + radiotherapy (9
Gy .times. 3) + cyclophosphamide
[0256] A DLT is defined as any of the following: a non-hematologic
toxicity (e.g., uveitis, or any other irAE), or a hematologic
toxicity (e.g., neutropenia, thrombocytopenia, febrile
neutropenia).
[0257] The maximum tolerated dose (MTD) is defined as the highest
dose at which fewer than a third of an expanded cohort of 6
patients experience a DLT during the first cycle of treatment.
Thus, the MTD is defined as the dose level immediately below the
level at which dosing is stopped due to the occurrence of 2 or more
DLTs in an expanded cohort of 6 patients. If dose escalation is not
stopped due to the occurrence of DLTs, it will be considered that
the MTD has not been determined. It is possible that an MTD may not
be defined in this study, either for a monotherapy group or for
individual combination groups. Additionally, it is possible that
mAb MTDs may differ between monotherapy and each combination
treatment regimen.
Study Duration
[0258] Patients will receive up to 48 weeks of treatment, after
which there will be a 24 week follow-up period. A patient will
receive treatment until the 48 week treatment period is complete,
or until disease progression, unacceptable toxicity, withdrawal of
consent, or meeting of another study withdrawal criterion. After a
minimum of 24 weeks of treatment, patients with confirmed complete
responses (CR) may elect to discontinue treatment and continue with
all relevant study assessments (e.g., efficacy assessments). After
a minimum of 24 weeks of treatment, patients with tumor burden
assessments of stable disease (SD) or partial response (PR) that
have been unchanged for 3 successive tumor evaluations may also
elect to discontinue treatment and continue with all relevant study
assessments (e.g., efficacy assessments).
Study Population
[0259] The target population for this study comprises patients with
advanced malignancies who are not candidates for standard therapy,
unwilling to undergo standard therapy, or for whom no available
therapy is expected to convey clinical benefit; and patients with
malignancies that are incurable and have failed to respond to or
showed tumor progression despite standard therapy.
[0260] Inclusion criteria: A patient must meet with the following
criteria to be eligible for inclusion in the study: (1)
demonstrated progression of a solid tumor with no alternative
standard-of-care therapeutic option available; (2) at least 1
lesion for response assessment. Patients assigned to radiotherapy
require at least one additional lesion that can be safely
irradiated while sparing the index lesions and for which radiation
at the limited, palliative doses contemplated would be considered
medically appropriate; (3) Eastern Cooperative Oncology Group
(ECOG) performance status .ltoreq.1; (4) more than 18 years old;
(5) hepatic function: a. total bilirubin .ltoreq.1.5.times. upper
limit of normal (ULN; if liver metastases .ltoreq.3.times. ULN), b.
transaminases .ltoreq.3.times. ULN (or .ltoreq.5.0.times. ULN, if
liver metastases), c. alkaline phosphatase (ALP) .ltoreq.2.5.times.
ULN (or 5.0.times. ULN, if liver metastases); (6) renal function:
serum creatinine .ltoreq.1.5.times. ULN; (7) neutrophil count (ANC)
.gtoreq.1.5.times.10.sup.9/L, c. platelet count
.gtoreq.75.times.10.sup.9/L; (8) ability to provide signed informed
consent; and (9) ability and willingness to comply with scheduled
visits, treatment plans, laboratory tests, and other study-related
procedures.
[0261] Exclusion criteria: A patient who meets any of the following
criteria will be excluded from the study: (1) Ongoing or recent
(within 5 years) evidence of significant autoimmune disease that
required treatment with systemic immunosuppressive treatments,
which may suggest risk for irAEs; (2) Prior treatment with an agent
that blocks the PD-1/PD-L1 pathway; (3) Prior treatment with other
immune modulating agents within fewer than 4 weeks or 4 half-lives,
whichever is greater, prior to the first dose of mAb; (4) Examples
of immune modulating agents include blockers of CTLA-4, 4-1BB
(CD137), OX-40, therapeutic vaccines, or cytokine treatments; (5)
Untreated brain metastasis(es) that may be considered active.
Patients with previously treated brain metastases may participate
provided they are stable (i.e., without evidence of progression by
imaging for at least 4 weeks prior to the first dose of study
treatment, and any neurologic symptoms have returned to baseline),
and there is no evidence of new or enlarging brain metastases; (6)
Immunosuppressive corticosteroid doses (>10 mg prednisone daily
or equivalent) within 4 weeks prior to the first dose of mAb; (7)
Deep vein thrombosis, pulmonary embolism (including asymptomatic
pulmonary embolism identified on imaging), or other thromboembolic
event within the 6 months preceding the first dose of mAb; (8)
Active infection requiring therapy, including known infection with
human immunodeficiency virus, or active infection with hepatitis B
or hepatitis C virus; (9) History of pneumonitis within the last 5
years; (10) Any investigational or antitumor treatment within 30
days prior to the initial administration of mAb; (11) History of
documented allergic reactions or acute hypersensitivity reaction
attributed to treatment with antibody therapies in general, or to
agents specifically used in the study; (12) Known allergy to
doxycycline or tetracycline (precaution due to presence of trace
components in mAb); (13) Breast-feeding; (14) Positive serum
pregnancy test; (15) History within the last 5 years of an invasive
malignancy other than the one treated in this study, with the
exception of resected/ablated basal or squamous-cell carcinoma of
the skin or carcinoma in situ of the cervix, or other local tumors
considered cured by local treatment; (16) Acute or chronic
psychiatric problems that, under the evaluation of the
investigator, make the patient ineligible for participation; and
(17) Continued sexual activity in men or women of childbearing
potential who are unwilling to practice adequate contraception
during the study.
Study Treatments
[0262] mAb will be supplied as a liquid in sterile, single-use
vials. Each vial will contain a volume sufficient to withdraw 10 mL
of mAb at a concentration of 25 mg/mL. Instructions on dose
preparation are provided in the study reference manuals. mAb will
be administered in an outpatient setting as a 30 minute IV
infusion. Each patient's dose will depend on individual body
weight. The dose of mAb must be adjusted each cycle for changes in
body weight of 0%. mAb will be administered alone and in
combination with radiation and or cyclophosphamide.
Monotherapy
[0263] mAb will be administered in an outpatient setting by IV
infusion over 30 minutes every 14 days for 48 weeks (i.e., Days 1,
15.+-.3, 29.+-.3, and 43.+-.3 of a 56 day cycle). Planned
monotherapy regimens to be assigned may include: (i) 1 mg/kg IV
infusion over 30 minutes every 14 days for 48 weeks; (ii) 3 mg/kg
infusion over 30 minutes every 14 days for 48 weeks; (iii) 10 mg/kg
infusion over 30 minutes every 14 days for 48 weeks; and (iv) 0.3
mg/kg infusion over 30 minutes every 14 days for 48 weeks (if MTD
is determined to be below 1 mg/kg).
Combination Therapy
[0264] Concomitant radiation therapy and cyclophosphamide will be
supplied through a prescription and their usage, dose, dose
modifications, reductions, or delays, as well as any potential AEs
resulting from their use, will be tracked along with that of
mAb.
[0265] Co-administration of mAb and radiation: mAb will be
administered by IV infusion over 30 minutes every 14 days for 48
weeks in combination with radiation treatment from day 8 to day 12.
Planned combination mAb and radiation therapy regimens may include:
[0266] 1 mg/kg mAb infusion over 30 minutes every 14 days for 48
weeks plus 30 Gy radiotherapy (6 Gy.times.5 times/week; given 1
week after the first dose of mAb, preferably on consecutive days)
[0267] 1 mg/kg mAb infusion over 30 minutes every 14 days for 48
weeks plus 27 Gy radiotherapy (9 Gy.times.3 times/week; given 1
week after the first dose of mAb, preferably not on consecutive
days) [0268] 3 mg/kg mAb infusion over 30 minutes every 14 days for
48 weeks plus 30 Gy radiotherapy (6 Gy.times.5 times/week; given 1
week after the first dose of mAb, preferably on consecutive days)
[0269] 3 mg/kg mAb infusion over 30 minutes every 14 days for 48
weeks plus 27 Gy radiotherapy (9 Gy.times.3 times/week; given 1
week after the first dose of mAb, preferably not on consecutive
days)
[0270] Patients will receive either 30 Gy given as 5 fractions of 6
Gy administered daily starting 1 week after the first dose of mAb,
or 27 Gy given as 3 fractions of 9 Gy administered every other day
starting 1 week after the first dose of mAb. The lesion selected
for radiation should be a lesion that can be safely irradiated with
focal irradiation while sparing the index lesion(s), and for which
radiation at the limited, palliative doses contemplated would be
considered medically appropriate. The target dose for a patient
will be based on cohort assignment and should conform to the normal
tissue requirements, in accord with standard radiation oncology
practice. Treatment at the protocol-specified dosing regimen is
permitted only if the normal tissue criteria are met. If the normal
tissue criteria cannot be met at either of the radiation therapy
regiments specified in the protocol, the patient is not eligible
for enrollment in a combination radiation treatment cohort in this
study.
[0271] Co-administration of mAb and cyclophosphamide: mAb will be
administered by IV infusion over 30 minutes every 14 days (2 weeks)
for 48 weeks in combination with cyclophosphamide 200 mg/m.sup.2
every 14 days for 4 doses. Each of the 4 cyclophosphamide doses
will be administered 1 day before each of the first 4 mAb doses
(days -1, 14, 28, and 42 of the first 56 day cycle).
[0272] Though cyclophosphamide has been used successfully
concurrently with other drugs, the rate of metabolism and the
leukopenic activity of cyclophosphamide reportedly are increased by
chronic administration of high doses of phenobarbital.
Cyclophosphamide treatment causes a marked and persistent
inhibition of cholinesterase activity, thus potentiating the effect
of succinylcholine chloride. The planned combination mAb and
cyclophosphamide regimen to be assigned is: [0273] Cyclophosphamide
200 mg/m.sup.2 every 14 days (days -1, 14, 28, and 42 of the first
56 day cycle) for a total of 4 doses plus [0274] 3 mg/kg mAb
infusion over 30 minutes every 14 days for 48 weeks (provided
monotherapy dose of 3 mg/kg<MTD; if 3 mg/kg>MTD, dose will be
1 mg/kg).
[0275] Co-administration of mAb, radiation and cyclophosphamide:
The planned combination mAb, radiation, and cyclophosphamide
regimen includes: [0276] Cyclophosphamide 200 mg/m.sup.2 every 14
days (days -1, 14, 28, and 42 of the first 56 day cycle) for a
total of 4 doses [0277] plus [0278] 27 Gy radiotherapy (9
Gy.times.3 times/week; given 1 week after the first dose of mAb,
preferably not on consecutive days) OR [0279] 30 Gy radiotherapy (6
Gy.times.5 times/week; given 1 week after the first dose of mAb,
preferably on consecutive days) [0280] plus [0281] 3 mg/kg mAb
infusion over 30 minutes every 14 days for 48 weeks (provided
monotherapy dose of 3 mg/kg<MTD; if 3 mg/kg>MTD, dose will be
1 mg/kg)
Study Variables
[0282] Primary Variables: Primary safety variables include
incidence of DLTs, incidence and severity of treatment-emergent
adverse events (TEAEs), and abnormal laboratory findings through 48
weeks of treatment.
[0283] Secondary Variables: Key secondary variables include the
following: [0284] Serum concentration and pharmacokinetics (PK) of
mAb [0285] Antitumor activities assessed using the appropriate
criteria for the indication: [0286] Response Evaluation Criteria in
Solid Tumors (RECIST) criteria measured by computed tomography (CT)
or magnetic resonance imaging (MRI) [0287] Other assessment
criteria should also be used for specific tumors in which RECIST
measurements are not the standard. [0288] Immune-Related Response
Criteria (irRC) applied to RECIST measurements. [0289] In all
cases, irRC will be the governing tool to determine progression of
disease (PD), SD, CR, or PR. Standard RECIST data will also be
collected for information purposes. [0290] Anti-mAb antibodies
Study Procedures
[0291] The following procedures will be performed at screening for
the purpose of determining study eligibility or characterizing the
baseline population: (i) serum .beta.-HCG (result must be
.ltoreq.72 hours before first dose); (ii) Collection of archived
tumor material: After a patient has given informed consent, the
patient will be asked to arrange to provide any available
previously collected tumor samples; (iii) Brain MRI: Brain MRI is
required at screening if not performed in the prior 60 days; and
(iv) Chest x-ray: Chest is x-ray required at screening if not
performed in the prior 60 days.
[0292] Efficacy Procedures: A CT or MRI for tumor assessment will
be performed at the screening visit (within 28 days prior to
infusion) and during every cycle (approximately every 8 weeks) on
day 56.+-.3, and when disease progression is suspected.
Additionally, for patients who have not progressed on study, tumor
assessment will be performed for follow-up visits 3, 5, and 7. Once
the choice has been made to use CT scan or MRI, subsequent
assessments will be made using the same modality.
[0293] Tumor response evaluation will be performed according to
immune-related response criteria (irRC; Nishino 2013). Assessments
according to Response Evaluation Criteria in Solid Tumors (RECIST)
version 1.1 (Eisenhauer 2009) will also be performed as a
supportive exploration; however, the primary determination of
disease progression for an individual patient will be made
according to irRC. Measurable lesions selected as target lesions
for RECIST assessments will also be included as index lesions for
irRC assessments.
[0294] Safety Procedures: Vital signs, including temperature,
resting blood pressure, pulse, and respiration, will be collected.
When scheduled at the same visit as other procedures, vital signs
should be measured prior to clinical laboratory assessments, PK, or
exploratory sample collection. During cycle 1, vital signs will be
recorded on treatment days prior to treatment, at the end of the
infusion, every 30 minutes for the first 4 hours post-infusion, and
at 6 and 8 hours post study drug administration. On subsequent
cycles, vital signs on treatment days will be assessed and
documented prior to the infusion, every 30 minutes for the first 2
hours, and then hourly until 4 hours following study drug
administration.
[0295] A thorough complete or limited physical examination will be
performed at visits. Complete physical examination will include
examination of skin, head, eyes, nose, throat, neck, joints, lungs,
heart, pulse, abdomen (including liver and spleen), lymph nodes,
and extremities, as well as a brief neurologic examination. Limited
physical examination will include lungs, heart, abdomen, and
skin.
[0296] A standard 12-lead ECG will be performed. Any ECG finding
that is judged by the investigator as a clinically significant
change (worsening) compared to the baseline value will be
considered an AE, recorded, and monitored.
[0297] Immune safety assays consist of rheumatoid factor (RF),
thyroid stimulating hormone (TSH), C-reactive protein (CRP), and
antinuclear antibody (ANA) titer and pattern. If, during the course
of the study, a 4-fold or greater increase from baseline in RF or
ANA or abnormal levels of TSH or CRP are observed, the following
tests may also be performed: anti-DNA antibody, anti-Sjogren's
syndrome A antigen (SSA) antibody (Ro), anti-Sjogren's syndrome B
antigen (SSB) antibody (La), antithyroglobulin antibody, anti-LKM
antibody, antiphospholipid antibody, anti-islet cell antibody,
antineutrophil cytoplasm antibody, C3, C4, CH50. Activated partial
thromboplastin time (aPTT) and International Normalized Ratio (INR)
will be analyzed by the site's local laboratory.
Safety
[0298] An adverse event (AE) is any untoward medical occurrence in
a patient administered a study drug which may or may not have a
causal relationship with the study drug. Therefore, an AE is any
unfavorable and unintended sign (including abnormal laboratory
finding), symptom, or disease which is temporally associated with
the use of a study drug, whether or not considered related to the
study drug. An AE also includes any worsening (i.e., any clinically
significant change in frequency and/or intensity) of a pre-existing
condition that is temporally associated with the use of the study
drug. Progression of underlying malignancy will not be considered
an AE if it is clearly consistent with the typical progression
pattern of the underlying cancer (including time course, affected
organs, etc.). Clinical symptoms of progression may be reported as
AEs if the symptom cannot be determined as exclusively due to the
progression of the underlying malignancy, or does not fit the
expected pattern of progression for the disease under study.
[0299] An serious adverse event (SAE) is any untoward medical
occurrence that at any dose results in death, is life-threatening,
requires in-patient hospitalization or prolongation of existing
hospitalization, results in persistent or significant
disability/incapacity (substantial disruption of one's ability to
conduct normal life functions), is a congenital anomaly/birth
defect.
[0300] Patient information on all AEs and SAEs will be
recorded.
Statistical Plan
[0301] The study dose escalation is based on a traditional 3+3
design with 3 to 6 patients assigned per dose level. The exact
number of patients enrolled in the study will depend on the number
of protocol-defined DLTs observed, and the need to expand currently
defined dose levels, or open additional cohorts at lower dose
levels. After the required initial enrollment to the next cohort in
the dose escalation has occurred, enrollment to each of the
previous cohorts below the MTD for that treatment will be expanded
(if not previously expanded during escalation) to a total of 6
patients.
[0302] Data will be summarized using descriptive statistics only.
In general, data will be summarized by dose levels and
combinations. The safety summaries and analyses will be performed
on the safety analysis set (SAF). The primary analysis of safety
will be based on treatment-emergent AEs (TEAEs).
[0303] The present invention is not to be limited in scope by the
specific embodiments described herein. Indeed, various
modifications of the invention in addition to those described
herein will become apparent to those skilled in the art from the
foregoing description and the accompanying figures. Such
modifications are intended to fall within the scope of the appended
claims.
Sequence CWU 1 SEQUENCE LISTING <160> NUMBER OF SEQ ID
NOS: 337 <210> SEQ ID NO 1 <211> LENGTH: 363
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 1 gaggtgcagc tgttggagtc tgggggaggc ttggtacagc
ctggggggtc cctgagactc 60 tcctgttcag cctctggatt cacctttagc
agctatacca tgaactgggt ccgccaggct 120 ccagggaagg ggctggagtg
ggtctcaggt attagtgata ccggtggtaa cacatactac 180 acagactccg
tgaagggccg gttcaccgtc tccagagaca attccaagaa cacactgtct 240
ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcag
300 ggtggaagtt acccctatta ctttcactac tggggccagg gatccctggt
caccgtctcc 360 tca 363 <210> SEQ ID NO 2 <211> LENGTH:
121 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 2 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu
Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ser Ala Ser
Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Thr Met Asn Trp Val Arg Gln
Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Asp
Thr Gly Gly Asn Thr Tyr Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg
Phe Thr Val Ser Arg Asp Asn Ser Lys Asn Thr Leu Ser 65 70 75 80 Leu
Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90
95 Ala Lys Asp Gln Gly Gly Ser Tyr Pro Tyr Tyr Phe His Tyr Trp Gly
100 105 110 Gln Gly Ser Leu Val Thr Val Ser Ser 115 120 <210>
SEQ ID NO 3 <211> LENGTH: 24 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 3
ggattcacct ttagcagcta tacc 24 <210> SEQ ID NO 4 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 4 Gly Phe Thr Phe Ser Ser Tyr Thr 1
5 <210> SEQ ID NO 5 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 5
attagtgata ccggtggtaa caca 24 <210> SEQ ID NO 6 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 6 Ile Ser Asp Thr Gly Gly Asn Thr 1
5 <210> SEQ ID NO 7 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 7
gcgaaagatc agggtggaag ttacccctat tactttcact ac 42 <210> SEQ
ID NO 8 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 8 Ala Lys Asp
Gln Gly Gly Ser Tyr Pro Tyr Tyr Phe His Tyr 1 5 10 <210> SEQ
ID NO 9 <211> LENGTH: 324 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 9 gacatccaga
tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60
atcacttgcc gggcaagtca gagcattagc agctatttaa tttggtatca gcagaaacca
120 gggacagccc ctaagttcct gatctatgct gcatccagtt tgcaaagtgg
ggtcccatca 180 aggttcagtg gctgtggatc tgggacagat ttcactctca
ccatcagcag tctgcaacct 240 gaagattttg caacttacta ctgtcaacag
agttacagta cccctccgat caccttcggc 300 caagggacac gactggagat taaa 324
<210> SEQ ID NO 10 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 10
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser
Tyr 20 25 30 Leu Ile Trp Tyr Gln Gln Lys Pro Gly Thr Ala Pro Lys
Phe Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Cys Gly Ser Gly Thr Asp Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln
Gly Thr Arg Leu Glu Ile Lys 100 105 <210> SEQ ID NO 11
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 11 cagagcatta gcagctat
18 <210> SEQ ID NO 12 <211> LENGTH: 6 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 12
Gln Ser Ile Ser Ser Tyr 1 5 <210> SEQ ID NO 13 <211>
LENGTH: 9 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 13 gctgcatcc 9 <210> SEQ ID
NO 14 <211> LENGTH: 3 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 14 Ala Ala Ser 1
<210> SEQ ID NO 15 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 15
caacagagtt acagtacccc tccgatcacc 30 <210> SEQ ID NO 16
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 16 Gln Gln Ser Tyr Ser
Thr Pro Pro Ile Thr 1 5 10 <210> SEQ ID NO 17 <211>
LENGTH: 366 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 17 gaggtgcagc tggtggagtc tggaggaggc
ttggtccagc ctggggggtc cctgagactc 60 tcctgtgcag cctctgggtt
caccgtcagt aacaactaca tgagctgggt ccgccaggct 120 ccagggaagg
ggctggagtg ggtctcagtt atttatagcg gtggtttcac atactacaca 180
gactccgtga agggccgatt caccatctcc agacacaatt ccaagaacac gctgtatctt
240 caaatgaaca gcctgagagc tgaggacacg gccgtgtatt actgtgcgag
gtattactat 300 gatactagtg attattggac cttctttgac tactggggcc
agggaaccct ggtcaccgtc 360 tcctca 366 <210> SEQ ID NO 18
<211> LENGTH: 122 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 18 Glu Val Gln Leu Val
Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg
Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Asn Asn 20 25 30 Tyr
Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40
45 Ser Val Ile Tyr Ser Gly Gly Phe Thr Tyr Tyr Thr Asp Ser Val Lys
50 55 60 Gly Arg Phe Thr Ile Ser Arg His Asn Ser Lys Asn Thr Leu
Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val
Tyr Tyr Cys Ala 85 90 95 Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp
Thr Phe Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 19 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 19 gggttcaccg tcagtaacaa ctac 24 <210>
SEQ ID NO 20 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 20
Gly Phe Thr Val Ser Asn Asn Tyr 1 5 <210> SEQ ID NO 21
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 21 atttatagcg
gtggtttcac a 21 <210> SEQ ID NO 22 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 22 Ile Tyr Ser Gly Gly Phe Thr 1 5
<210> SEQ ID NO 23 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 23
gcgaggtatt actatgatac tagtgattat tggaccttct ttgactac 48 <210>
SEQ ID NO 24 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 24
Ala Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp Thr Phe Phe Asp Tyr 1 5
10 15 <210> SEQ ID NO 25 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 25 gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga
aagagccacc 60 ctctcctgca gggccagtca gagtgttagc agcaacttag
cctggtacca gcagaaacct 120 ggccaggctc ccaggctcct catctatggt
gcatccacca gggccactgg tatcccagcc 180 aggttcagtg gcagtgggtc
tgggacagag ttcactctca ccatcagtag cctgcagtct 240 ggagattttg
cagtttatta ctgtcagcag tataataact ggccgctcac tttcggcgga 300
gggaccaagg tggagatcaa t 321 <210> SEQ ID NO 26 <211>
LENGTH: 107 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 26 Glu Ile Val Met Thr Gln Ser Pro
Ala Thr Leu Ser Val Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Asn 20 25 30 Leu Ala Trp Tyr
Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Gly
Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser 65
70 75 80 Gly Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn Asn Trp
Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Asn 100
105 <210> SEQ ID NO 27 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 27 cagagtgtta gcagcaac 18 <210> SEQ ID NO 28
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 28 Gln Ser Val Ser Ser
Asn 1 5 <210> SEQ ID NO 29 <211> LENGTH: 9 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 29 ggtgcatcc 9 <210> SEQ ID NO 30 <211>
LENGTH: 3 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 30 Gly Ala Ser 1 <210> SEQ ID
NO 31 <211> LENGTH: 27 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 31 cagcagtata
ataactggcc gctcact 27 <210> SEQ ID NO 32 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 32 Gln Gln Tyr Asn Asn Trp Pro Leu Thr 1 5
<210> SEQ ID NO 33 <211> LENGTH: 366 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 33
gaggtgcagc tggtggagtc tggaggaggc ttggtccagc ctggggggtc cctgagactc
60 tcctgtgcag cctctgggtt caccgtcagt aacaactaca tgagctgggt
ccgccaggct 120 ccagggaagg ggctggagtg ggtctcagtt atttatagcg
gtggtttcac atactacaca 180 gactccgtga agggccgatt caccatctcc
agacacaatt ccaagaacac gctgtatctt 240 caaatgaaca gcctgagagc
tgaggacacg gccgtgtatt actgtgcgag gtattactat 300 gatactagtg
attattggac cttctttgac tactggggcc agggaaccct ggtcaccgtc 360 tcctca
366 <210> SEQ ID NO 34 <211> LENGTH: 122 <212>
TYPE: PRT <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 34 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln
Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe
Thr Val Ser Asn Asn 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro
Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Val Ile Tyr Ser Gly Gly
Phe Thr Tyr Tyr Thr Asp Ser Val Lys 50 55 60 Gly Arg Phe Thr Ile
Ser Arg His Asn Ser Lys Asn Thr Leu Tyr Leu 65 70 75 80 Gln Met Asn
Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala 85 90 95 Arg
Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp Thr Phe Phe Asp Tyr Trp 100 105
110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> SEQ
ID NO 35 <211> LENGTH: 24 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 35 gggttcaccg
tcagtaacaa ctac 24 <210> SEQ ID NO 36 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 36 Gly Phe Thr Val Ser Asn Asn Tyr 1 5
<210> SEQ ID NO 37 <211> LENGTH: 21 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 37
atttatagcg gtggtttcac a 21 <210> SEQ ID NO 38 <211>
LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 38 Ile Tyr Ser Gly Gly Phe Thr 1 5
<210> SEQ ID NO 39 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 39
gcgaggtatt actatgatac tagtgattat tggaccttct ttgactac 48 <210>
SEQ ID NO 40 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 40
Ala Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp Thr Phe Phe Asp Tyr 1 5
10 15 <210> SEQ ID NO 41 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 41 gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga
aagagccacc 60 ctctcctgca gggccagtca gagtgttagc agcaacttag
cctggtacca gcagaaacct 120 ggccaggctc ccaggctcct catctatggt
gcatccacca gggccactgg tatcccagcc 180 aggttcagtg gcagtgggtc
tgggacagag ttcactctca ccatcagtag cctgcagtct 240 ggagattttg
cagtttatta ctgtcagcag tataataact ggccgctcac tttcggcgga 300
gggaccaagg tggagatcaa t 321 <210> SEQ ID NO 42 <211>
LENGTH: 107 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 42 Glu Ile Val Met Thr Gln Ser Pro
Ala Thr Leu Ser Val Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Asn 20 25 30 Leu Ala Trp Tyr
Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Gly
Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser 65
70 75 80 Gly Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn Asn Trp
Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Asn 100
105 <210> SEQ ID NO 43 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 43 cagagtgtta gcagcaac 18 <210> SEQ ID NO 44
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 44 Gln Ser Val Ser Ser
Asn 1 5 <210> SEQ ID NO 45 <211> LENGTH: 9 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 45 ggtgcatcc 9 <210> SEQ ID NO 46 <211>
LENGTH: 3 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 46 Gly Ala Ser 1 <210> SEQ ID
NO 47 <211> LENGTH: 27 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 47 cagcagtata
ataactggcc gctcact 27 <210> SEQ ID NO 48 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 48 Gln Gln Tyr Asn Asn Trp Pro Leu Thr 1 5
<210> SEQ ID NO 49 <211> LENGTH: 357 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 49
caggtgcagc tggtggagtc tgggggaggc gtggtccagt ctgggaggtc cctgagactc
60 tcctgtgcag cgtctggatt caccttcagt agctatggca tgcactgggt
ccgccaggct 120 ccaggcaagg ggctggagtg ggtggcagtt atatggtatg
atggaagtaa tatatactat 180 tcagactccg tgaagggccg attcaccatc
tccagagcca attccaagaa cacgctgtat 240 ctgcaaatga acagcctgag
agccgaggac acggctgttt attactgtgc gagaccggga 300 cactggaact
acttctttga atactggggc cagggaaccc tggtcaccgt ctcctca 357 <210>
SEQ ID NO 50 <211> LENGTH: 119 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 50
Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Ser Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser
Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ala Val Ile Trp Tyr Asp Gly Ser Asn Ile Tyr
Tyr Ser Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Ala
Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Pro Gly His
Trp Asn Tyr Phe Phe Glu Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val
Thr Val Ser Ser 115 <210> SEQ ID NO 51 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 51 ggattcacct tcagtagcta tggc 24 <210>
SEQ ID NO 52 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 52
Gly Phe Thr Phe Ser Ser Tyr Gly 1 5 <210> SEQ ID NO 53
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 53 atatggtatg
atggaagtaa tata 24 <210> SEQ ID NO 54 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 54 Ile Trp Tyr Asp Gly Ser Asn Ile 1 5
<210> SEQ ID NO 55 <211> LENGTH: 36 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 55
gcgagaccgg gacactggaa ctacttcttt gaatac 36 <210> SEQ ID NO 56
<211> LENGTH: 12 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 56 Ala Arg Pro Gly His
Trp Asn Tyr Phe Phe Glu Tyr 1 5 10 <210> SEQ ID NO 57
<211> LENGTH: 324 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 57 gacatccaga
tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60
atcacttgcc gggcaagtca gagcattaac aactatttaa attggtatca gcagaaacca
120 gggaaagccc ctaagctcct gatctatact gcatccagtt tgcaaagtgg
ggtcccatca 180 aggttcagtg gcagtggatc tgggacagat ttcactctca
ccatcagcag tctgcaacct 240 gaagattttg caacttacta ctgtcaacag
agttacagta cccctccgct caccttcggc 300 caagggacac aactggagat taaa 324
<210> SEQ ID NO 58 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 58
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Asn Asn
Tyr 20 25 30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45 Tyr Thr Ala Ser Ser Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Leu Thr Phe Gly Gln
Gly Thr Gln Leu Glu Ile Lys 100 105 <210> SEQ ID NO 59
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 59 cagagcatta acaactat
18 <210> SEQ ID NO 60 <211> LENGTH: 6 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 60
Gln Ser Ile Asn Asn Tyr 1 5 <210> SEQ ID NO 61 <211>
LENGTH: 9 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 61 actgcatcc 9 <210> SEQ ID
NO 62 <211> LENGTH: 3 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 62 Thr Ala Ser 1
<210> SEQ ID NO 63 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 63
caacagagtt acagtacccc tccgctcacc 30 <210> SEQ ID NO 64
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 64 Gln Gln Ser Tyr Ser
Thr Pro Pro Leu Thr 1 5 10 <210> SEQ ID NO 65 <211>
LENGTH: 366 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 65 gaggtgcagc tggtggagtc tgggggaggc
ttggtacagc ctggggggtc cctgagactc 60 tcctgtggag cctctggatt
caccttcagg aactacgaca tgcactgggt ccgccaaatt 120 acaggaaaag
gtctggagtg ggtctcagct attggtagtg ctggtgacac atactatcca 180
gactccgtga agggccgatt caccatctcc agagaaaatg ccaagaactc cttgtatctt
240 caaatgaaca gcctgagagt cggggacacg gctgtgtatt actgtacaag
agatatccat 300 tgtagtagta ccaggtgcta cggtatggac gtctggggcc
aagggaccac ggtcaccgtc 360 tcctca 366 <210> SEQ ID NO 66
<211> LENGTH: 122 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 66 Glu Val Gln Leu Val
Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg
Leu Ser Cys Gly Ala Ser Gly Phe Thr Phe Arg Asn Tyr 20 25 30 Asp
Met His Trp Val Arg Gln Ile Thr Gly Lys Gly Leu Glu Trp Val 35 40
45 Ser Ala Ile Gly Ser Ala Gly Asp Thr Tyr Tyr Pro Asp Ser Val Lys
50 55 60 Gly Arg Phe Thr Ile Ser Arg Glu Asn Ala Lys Asn Ser Leu
Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Val Gly Asp Thr Ala Val
Tyr Tyr Cys Thr 85 90 95 Arg Asp Ile His Cys Ser Ser Thr Arg Cys
Tyr Gly Met Asp Val Trp 100 105 110 Gly Gln Gly Thr Thr Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 67 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 67 ggattcacct tcaggaacta cgac 24 <210>
SEQ ID NO 68 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 68
Gly Phe Thr Phe Arg Asn Tyr Asp 1 5 <210> SEQ ID NO 69
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 69 attggtagtg
ctggtgacac a 21 <210> SEQ ID NO 70 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 70 Ile Gly Ser Ala Gly Asp Thr 1 5
<210> SEQ ID NO 71 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 71
acaagagata tccattgtag tagtaccagg tgctacggta tggacgtc 48 <210>
SEQ ID NO 72 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 72
Thr Arg Asp Ile His Cys Ser Ser Thr Arg Cys Tyr Gly Met Asp Val 1 5
10 15 <210> SEQ ID NO 73 <211> LENGTH: 324 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 73 gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga
cagagtcacc 60 atcacttgcc gggcaagtca gagcattagc aactatttaa
attggtatca gcagaaacca 120 gggaaagccc ctaagctcct gatctatgct
gcatccagtt tgcaaagtgg ggtcccatca 180 aggttcagtg gcagtggatc
tgggacagat ttcactctca ccatcagcag tctgcaacct 240 gaagattttg
caacttacta ctgtcaacag agttacagta cccctccgat caccttcggc 300
caagggacac gactggagat taaa 324 <210> SEQ ID NO 74 <211>
LENGTH: 108 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 74 Asp Ile Gln Met Thr Gln Ser Pro
Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr
Cys Arg Ala Ser Gln Ser Ile Ser Asn Tyr 20 25 30 Leu Asn Trp Tyr
Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Ala
Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65
70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr
Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys
100 105 <210> SEQ ID NO 75 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 75 cagagcatta gcaactat 18 <210> SEQ ID NO 76
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 76 Gln Ser Ile Ser Asn
Tyr 1 5 <210> SEQ ID NO 77 <211> LENGTH: 9 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 77 gctgcatcc 9 <210> SEQ ID NO 78 <211>
LENGTH: 3 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 78 Ala Ala Ser 1 <210> SEQ ID
NO 79 <211> LENGTH: 30 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 79 caacagagtt
acagtacccc tccgatcacc 30 <210> SEQ ID NO 80 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 80 Gln Gln Ser Tyr Ser Thr Pro Pro
Ile Thr 1 5 10 <210> SEQ ID NO 81 <211> LENGTH: 390
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 81 gaggtgcagc tggtggagtc tgggggaggc
ttggtaaagc ctggggggtc ccttcgactc 60 tcctgtgcag cctctggatt
caaattcagt aatgaatgga tgagctgggt ccgccaggct 120 ccagggaagg
ggctggagtg ggttggccgt attaaaagca aaactgatgg tgggacaaca 180
gactacgctg cacccgtgaa aggcagattc accatctcaa gagatgattc aaaaaatacg
240 ctgtatctgc aaatgaacag cctgaaaacc gaggacacag ccgtgtatta
ctgtaccaca 300 gatcaagatt tttggagtgg ttattatacc ggggctgact
actacggtat ggacgtctgg 360 ggccaaggga ccatggtcac cgtctcctca 390
<210> SEQ ID NO 82 <211> LENGTH: 130 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 82
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Lys Phe Ser Asn
Glu 20 25 30 Trp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Gly Arg Ile Lys Ser Lys Thr Asp Gly Gly Thr
Thr Asp Tyr Ala Ala 50 55 60 Pro Val Lys Gly Arg Phe Thr Ile Ser
Arg Asp Asp Ser Lys Asn Thr 65 70 75 80 Leu Tyr Leu Gln Met Asn Ser
Leu Lys Thr Glu Asp Thr Ala Val Tyr 85 90 95 Tyr Cys Thr Thr Asp
Gln Asp Phe Trp Ser Gly Tyr Tyr Thr Gly Ala 100 105 110 Asp Tyr Tyr
Gly Met Asp Val Trp Gly Gln Gly Thr Met Val Thr Val 115 120 125 Ser
Ser 130 <210> SEQ ID NO 83 <211> LENGTH: 24 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 83 ggattcaaat tcagtaatga atgg 24 <210> SEQ ID NO 84
<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 84 Gly Phe Lys Phe Ser
Asn Glu Trp 1 5 <210> SEQ ID NO 85 <211> LENGTH: 30
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 85 attaaaagca aaactgatgg tgggacaaca 30
<210> SEQ ID NO 86 <211> LENGTH: 10 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 86
Ile Lys Ser Lys Thr Asp Gly Gly Thr Thr 1 5 10 <210> SEQ ID
NO 87 <211> LENGTH: 63 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 87 accacagatc
aagatttttg gagtggttat tataccgggg ctgactacta cggtatggac 60 gtc 63
<210> SEQ ID NO 88 <211> LENGTH: 21 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 88
Thr Thr Asp Gln Asp Phe Trp Ser Gly Tyr Tyr Thr Gly Ala Asp Tyr 1 5
10 15 Tyr Gly Met Asp Val 20 <210> SEQ ID NO 89 <211>
LENGTH: 324 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 89 gacatccaga tgacccagtc tccatcctcc
ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgcc gggcaagtca
gagcattagc agctatttaa attggtatca gcagaaacca 120 gggaaagccc
ctaagctcct gatctatgct gcatccagtt tgcaaagtgg ggtcccatca 180
aggttcagtg gcagtggatc tgggacagat ttcactctca ccatcagcag tctgcaacct
240 gaagattttg caacttacta ctgtcaacag agttacagta cccctccgat
caccttcggc 300 caagggacac gactggagat taaa 324 <210> SEQ ID NO
90 <211> LENGTH: 108 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 90 Asp Ile Gln
Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp
Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr 20 25
30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile
35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe
Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser
Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln
Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly Thr Arg
Leu Glu Ile Lys 100 105 <210> SEQ ID NO 91 <211>
LENGTH: 18 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 91 cagagcatta gcagctat 18
<210> SEQ ID NO 92 <211> LENGTH: 6 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 92
Gln Ser Ile Ser Ser Tyr 1 5 <210> SEQ ID NO 93 <211>
LENGTH: 9 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 93 gctgcatcc 9 <210> SEQ ID
NO 94 <211> LENGTH: 3 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 94 Ala Ala Ser 1
<210> SEQ ID NO 95 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 95
caacagagtt acagtacccc tccgatcacc 30 <210> SEQ ID NO 96
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 96 Gln Gln Ser Tyr Ser
Thr Pro Pro Ile Thr 1 5 10 <210> SEQ ID NO 97 <211>
LENGTH: 348 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 97 cagatgcagc tccaacagtg gggcgcagga
ctattgaagc cttcggagac cctgtccctc 60 acctgcgttg tctatggtgg
gtccctcaat ggatactatt ggagctggat ccgccagtcc 120 cccgggaagg
ggctggagtg gattggggaa atcgatcata gtggaagcac caactacaac 180
ccgtccctca agaatcgagt caccatgtca gtagacacgt ctaagattca gttctccctg
240 aaactgacct ctgtgaccgt cgcggacacg gctgtgtatt actgtgcgag
agaaggatta 300 ttaccctttg actattgggg ccagggaacc ctggtcaccg tctcctca
348 <210> SEQ ID NO 98 <211> LENGTH: 116 <212>
TYPE: PRT <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 98 Gln Met Gln Leu Gln Gln Trp Gly Ala Gly Leu Leu Lys
Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Val Val Tyr Gly Gly
Ser Leu Asn Gly Tyr 20 25 30 Tyr Trp Ser Trp Ile Arg Gln Ser Pro
Gly Lys Gly Leu Glu Trp Ile 35 40 45 Gly Glu Ile Asp His Ser Gly
Ser Thr Asn Tyr Asn Pro Ser Leu Lys 50 55 60 Asn Arg Val Thr Met
Ser Val Asp Thr Ser Lys Ile Gln Phe Ser Leu 65 70 75 80 Lys Leu Thr
Ser Val Thr Val Ala Asp Thr Ala Val Tyr Tyr Cys Ala 85 90 95 Arg
Glu Gly Leu Leu Pro Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105
110 Thr Val Ser Ser 115 <210> SEQ ID NO 99 <211>
LENGTH: 24 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 99 ggtgggtccc tcaatggata ctat 24
<210> SEQ ID NO 100 <211> LENGTH: 8 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 100
Gly Gly Ser Leu Asn Gly Tyr Tyr 1 5 <210> SEQ ID NO 101
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 101 atcgatcata
gtggaagcac c 21 <210> SEQ ID NO 102 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 102 Ile Asp His Ser Gly Ser Thr 1 5
<210> SEQ ID NO 103 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 103
gcgagagaag gattattacc ctttgactat 30 <210> SEQ ID NO 104
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 104 Ala Arg Glu Gly
Leu Leu Pro Phe Asp Tyr 1 5 10 <210> SEQ ID NO 105
<211> LENGTH: 324 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 105 gaaattgtgt
tgacgcagtc tccaggcacc ctgtctttgt ctccagggga aagagtcacc 60
ctctcctgca gggccagtca gagtgtttac agcaactact tagcctggta ccagcagaat
120 cctggccagg ctcccaggct cctcatctat gctgcatcca acagggccac
tggcatccca 180 gacaggttca gtggcagtgg gtctgggaca gacttcactc
tcaccatcag cagactggag 240 cctgaagatt ttgcggtgta ttactgtcat
cagtatgcta cctcaccttg gacgttcggc 300 caagggacca aggtggaaat caaa 324
<210> SEQ ID NO 106 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 106
Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5
10 15 Glu Arg Val Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Tyr Ser
Asn 20 25 30 Tyr Leu Ala Trp Tyr Gln Gln Asn Pro Gly Gln Ala Pro
Arg Leu Leu 35 40 45 Ile Tyr Ala Ala Ser Asn Arg Ala Thr Gly Ile
Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr
Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr
Tyr Cys His Gln Tyr Ala Thr Ser Pro 85 90 95 Trp Thr Phe Gly Gln
Gly Thr Lys Val Glu Ile Lys 100 105 <210> SEQ ID NO 107
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 107 cagagtgttt
acagcaacta c 21 <210> SEQ ID NO 108 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 108 Gln Ser Val Tyr Ser Asn Tyr 1 5
<210> SEQ ID NO 109 <211> LENGTH: 9 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 109
gctgcatcc 9 <210> SEQ ID NO 110 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 110 Ala Ala Ser 1 <210> SEQ ID NO 111
<211> LENGTH: 27 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 111 catcagtatg
ctacctcacc ttggacg 27 <210> SEQ ID NO 112 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 112 His Gln Tyr Ala Thr Ser Pro Trp Thr 1 5
<210> SEQ ID NO 113 <211> LENGTH: 369 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 113
cagctgcagc tgcaggagtc gggcccagat ctggtgaagc cttcggatac cctgtccctc
60 acctgcactg tctctgatga ctccatcagc agtactactt actactgggc
ctggatccgc 120 cagcccccag ggaaggggct ggaatggatt ggcagtatgt
cttataatgg gaacaactac 180 tacaacccgt ccctcaagag tcgagtcgcc
atatccgcag gcacgtccca gaaacagttc 240 tccctgaaac tgacctctgt
gactgccgca gacacggctg tttatcactg tgcgagacat 300 cttggatata
acggcaactg gtaccccttt gacttctggg gccagggaat tctggtcacc 360
gtctcctct 369 <210> SEQ ID NO 114 <211> LENGTH: 123
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 114 Gln Leu Gln Leu Gln Glu Ser Gly Pro Asp
Leu Val Lys Pro Ser Asp 1 5 10 15 Thr Leu Ser Leu Thr Cys Thr Val
Ser Asp Asp Ser Ile Ser Ser Thr 20 25 30 Thr Tyr Tyr Trp Ala Trp
Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu 35 40 45 Trp Ile Gly Ser
Met Ser Tyr Asn Gly Asn Asn Tyr Tyr Asn Pro Ser 50 55 60 Leu Lys
Ser Arg Val Ala Ile Ser Ala Gly Thr Ser Gln Lys Gln Phe 65 70 75 80
Ser Leu Lys Leu Thr Ser Val Thr Ala Ala Asp Thr Ala Val Tyr His 85
90 95 Cys Ala Arg His Leu Gly Tyr Asn Gly Asn Trp Tyr Pro Phe Asp
Phe 100 105 110 Trp Gly Gln Gly Ile Leu Val Thr Val Ser Ser 115 120
<210> SEQ ID NO 115 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 115
gatgactcca tcagcagtac tacttactac 30 <210> SEQ ID NO 116
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 116 Asp Asp Ser Ile
Ser Ser Thr Thr Tyr Tyr 1 5 10 <210> SEQ ID NO 117
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 117 atgtcttata
atgggaacaa c 21 <210> SEQ ID NO 118 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 118 Met Ser Tyr Asn Gly Asn Asn 1 5
<210> SEQ ID NO 119 <211> LENGTH: 45 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 119
gcgagacatc ttggatataa cggcaactgg tacccctttg acttc 45 <210>
SEQ ID NO 120 <211> LENGTH: 15 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 120
Ala Arg His Leu Gly Tyr Asn Gly Asn Trp Tyr Pro Phe Asp Phe 1 5 10
15 <210> SEQ ID NO 121 <211> LENGTH: 324 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 121 gaaattgtgt tgacgcagtc tccaggcacc ctgtctttgt
ctccaggaga aagagccacc 60 ctctcctgca gggccagtca gagtgttagt
agtagttatt tagcctggta ccagcagaaa 120 cctggccagg ctcccaggct
cctcatctat ggtgcatcca gcaggaccac tggcatccca 180 gacaggttca
gtggcagtgg gtctgggaca gacttcactc tcaccatcag cagactggag 240
cctgaagatt ttgcagtgta ttactgtcag cagtatggta gctcaccttg gacgttcggc
300 caagggacca aggtggaaat caaa 324 <210> SEQ ID NO 122
<211> LENGTH: 108 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 122 Glu Ile Val Leu
Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg
Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser 20 25 30
Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35
40 45 Ile Tyr Gly Ala Ser Ser Arg Thr Thr Gly Ile Pro Asp Arg Phe
Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser
Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln
Tyr Gly Ser Ser Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr Lys Val
Glu Ile Lys 100 105 <210> SEQ ID NO 123 <211> LENGTH:
21 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 123 cagagtgtta gtagtagtta t 21 <210>
SEQ ID NO 124 <211> LENGTH: 7 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 124
Gln Ser Val Ser Ser Ser Tyr 1 5 <210> SEQ ID NO 125
<211> LENGTH: 9 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 125 ggtgcatcc 9
<210> SEQ ID NO 126 <211> LENGTH: 3 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 126
Gly Ala Ser 1 <210> SEQ ID NO 127 <211> LENGTH: 27
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 127 cagcagtatg gtagctcacc ttggacg 27
<210> SEQ ID NO 128 <211> LENGTH: 9 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 128
Gln Gln Tyr Gly Ser Ser Pro Trp Thr 1 5 <210> SEQ ID NO 129
<211> LENGTH: 366 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 129 gaagtgcagg
tggtagagtc tgggggcggc ttggtcgagc ctggcaggtc cctgagactc 60
tcctgtaaag cctctggatt cacctttgat gattatgcca tgcactgggt ccgacaaact
120 ccagggaagg ccctggagtg ggtctcgggt attaattgga gtggtaataa
cataggctat 180 gcggactctg tgaagggccg attcaccatc tccaaggacg
acgccaagaa ctccctgtat 240 ctgcaaatga acagtctgag acctgaggac
acggccttat attactgtac aaaagatata 300 agtataactg gaaccctcga
tgcttttgat gtctggggcc aagggacaat ggtcaccgtc 360 tcttca 366
<210> SEQ ID NO 130 <211> LENGTH: 122 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 130
Glu Val Gln Val Val Glu Ser Gly Gly Gly Leu Val Glu Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Lys Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Thr Pro Gly Lys Ala Leu
Glu Trp Val 35 40 45 Ser Gly Ile Asn Trp Ser Gly Asn Asn Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Lys Asp
Asp Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Pro Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Thr Lys Asp Ile Ser
Ile Thr Gly Thr Leu Asp Ala Phe Asp Val Trp 100 105 110 Gly Gln Gly
Thr Met Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 131
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 131 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 132 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 132 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 133 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 133
attaattgga gtggtaataa cata 24 <210> SEQ ID NO 134 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 134 Ile Asn Trp Ser Gly Asn Asn Ile
1 5 <210> SEQ ID NO 135 <211> LENGTH: 45 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 135 acaaaagata taagtataac tggaaccctc gatgcttttg atgtc 45
<210> SEQ ID NO 136 <211> LENGTH: 15 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 136
Thr Lys Asp Ile Ser Ile Thr Gly Thr Leu Asp Ala Phe Asp Val 1 5 10
15 <210> SEQ ID NO 137 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 137 gacatccaga tgacccagtc tccaatttcc gtgtctgcat
ctgtaggaga cagagtcacc 60 atcacttgtc gggcgagtca gggtattagc
aactggttag cctggtatca gcagaaacca 120 gggatagccc ctaaactcct
gatctattct gcatccagtt tacaaagtgg ggtcccatca 180 aggttcagag
gcagtggatc tgggacagac ttcactctca ccatcggcag cctgcagcct 240
gaagattttg caacttacta ttgtcaacag gctcacagtt tcccgctcac tttcggcgga
300 gggaccaagg tggagatcaa a 321 <210> SEQ ID NO 138
<211> LENGTH: 107 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 138 Asp Ile Gln Met
Thr Gln Ser Pro Ile Ser Val Ser Ala Ser Val Gly 1 5 10 15 Asp Arg
Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Asn Trp 20 25 30
Leu Ala Trp Tyr Gln Gln Lys Pro Gly Ile Ala Pro Lys Leu Leu Ile 35
40 45 Tyr Ser Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Arg
Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Gly Ser
Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ala
His Ser Phe Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu
Ile Lys 100 105 <210> SEQ ID NO 139 <211> LENGTH: 18
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 139 cagggtatta gcaactgg 18 <210> SEQ ID
NO 140 <211> LENGTH: 6 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 140 Gln Gly Ile
Ser Asn Trp 1 5 <210> SEQ ID NO 141 <211> LENGTH: 9
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 141 tctgcatcc 9 <210> SEQ ID NO 142
<211> LENGTH: 3 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 142 Ser Ala Ser 1
<210> SEQ ID NO 143 <211> LENGTH: 27 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 143
caacaggctc acagtttccc gctcact 27 <210> SEQ ID NO 144
<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 144 Gln Gln Ala His
Ser Phe Pro Leu Thr 1 5 <210> SEQ ID NO 145 <211>
LENGTH: 348 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 145 caggtgcaat tagtggagtc
tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60 tcctgtgcag
cgtctggatt caccttcagt agctatggca tgcactgggt ccgccaggct 120
ccaggcaagg ggctggagtg ggtggcaatt atatggtctg atggagatag tgaatataat
180 ctagactccg taaagggccg attcaccatc tccagagaca attccaagaa
cacgctgtat 240 ctgcaaatga acagtctgag agtcgaagac tcggctgtat
attactgtgc gagagatcga 300 gaccttgagg atatctgggg ccaagggaca
atggtcaccg tctcttca 348 <210> SEQ ID NO 146 <211>
LENGTH: 116 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 146 Gln Val Gln Leu Val Glu Ser Gly
Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys
Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Gly Met His Trp
Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ile
Ile Trp Ser Asp Gly Asp Ser Glu Tyr Asn Leu Asp Ser Val 50 55 60
Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65
70 75 80 Leu Gln Met Asn Ser Leu Arg Val Glu Asp Ser Ala Val Tyr
Tyr Cys 85 90 95 Ala Arg Asp Arg Asp Leu Glu Asp Ile Trp Gly Gln
Gly Thr Met Val 100 105 110 Thr Val Ser Ser 115 <210> SEQ ID
NO 147 <211> LENGTH: 24 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 147 ggattcacct
tcagtagcta tggc 24 <210> SEQ ID NO 148 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 148 Gly Phe Thr Phe Ser Ser Tyr Gly 1 5
<210> SEQ ID NO 149 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 149
atatggtctg atggagatag tgaa 24 <210> SEQ ID NO 150 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 150 Ile Trp Ser Asp Gly Asp Ser Glu
1 5 <210> SEQ ID NO 151 <211> LENGTH: 27 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 151 gcgagagatc gagaccttga ggatatc 27 <210> SEQ ID
NO 152 <211> LENGTH: 9 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 152 Ala Arg Asp
Arg Asp Leu Glu Asp Ile 1 5 <210> SEQ ID NO 153 <211>
LENGTH: 321 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 153 gacatccaga tgacccagtc
tccatcctcc ctgtctgcat ctgtcggaga cagagtcacc 60 atcacttgcc
gggcaagtca gggcattaga aatgatttag gctggtatca gcagaaacca 120
gggaaagccc ctaagcgcct gatctatgct gcatccaatt tgcaaagtgg ggtcccatca
180 aggttcagcg gcagtggatc tgggacagag ttcactctca caatcagcag
cctgcagcct 240 gaagattttg caacttatta ctgtctacag cataatagtt
atccgctcac tttcggcgga 300 gggaccaagg tggagatcaa a 321 <210>
SEQ ID NO 154 <211> LENGTH: 107 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 154
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Arg Asn
Asp 20 25 30 Leu Gly Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Arg Leu Ile 35 40 45 Tyr Ala Ala Ser Asn Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Leu Gln His Asn Ser Tyr Pro Leu 85 90 95 Thr Phe Gly Gly Gly
Thr Lys Val Glu Ile Lys 100 105 <210> SEQ ID NO 155
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 155 cagggcatta
gaaatgat 18 <210> SEQ ID NO 156 <211> LENGTH: 6
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 156 Gln Gly Ile Arg Asn Asp 1 5 <210>
SEQ ID NO 157 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 157
gctgcatcc 9 <210> SEQ ID NO 158 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 158 Ala Ala Ser 1 <210> SEQ ID NO 159
<211> LENGTH: 27 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 159 ctacagcata
atagttatcc gctcact 27 <210> SEQ ID NO 160 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 160 Leu Gln His Asn Ser Tyr Pro Leu Thr 1 5
<210> SEQ ID NO 161 <211> LENGTH: 351 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 161
gaggtgcagt tgttggagtc tgggggagtt ttggtacagc ctggggggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttagt aattttggca tgacgtgggt
ccgccaggct 120 ccagggaagg gactggagtg ggtctcaggt attagtggtg
gcggtcgtga cacatacttc 180 gcagactccg tgaagggccg gttcaccatc
tccagagaca attccaagaa tacgttgtat 240 ctacagatga acagcctgaa
aggcgaggac acggccgtat attactgtgt gaagtgggga 300 aatatttact
ttgactactg gggccaggga accctggtca ccgtctcatc a 351 <210> SEQ
ID NO 162 <211> LENGTH: 117 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 162 Glu Val Gln
Leu Leu Glu Ser Gly Gly Val Leu Val Gln Pro Gly Gly 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Phe 20 25
30 Gly Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ser Gly Ile Ser Gly Gly Gly Arg Asp Thr Tyr Phe Ala Asp
Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Gly Glu Asp
Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Trp Gly Asn Ile Tyr Phe
Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115
<210> SEQ ID NO 163 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 163
ggattcacct ttagtaattt tggc 24 <210> SEQ ID NO 164 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 164 Gly Phe Thr Phe Ser Asn Phe Gly
1 5 <210> SEQ ID NO 165 <211> LENGTH: 24 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 165 attagtggtg gcggtcgtga caca 24 <210> SEQ ID NO
166 <211> LENGTH: 8 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 166 Ile Ser Gly
Gly Gly Arg Asp Thr 1 5 <210> SEQ ID NO 167 <211>
LENGTH: 30 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 167 gtgaagtggg gaaatattta
ctttgactac 30 <210> SEQ ID NO 168 <211> LENGTH: 10
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 168 Val Lys Trp Gly Asn Ile Tyr Phe Asp Tyr 1
5 10 <210> SEQ ID NO 169 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 169 gacatccaga tgacccagtc tccatcctcc ctgtctgcat
ctgtcggaga cagcatcacc 60 atcacttgcc gggcgagtct gtccattaac
acctttttaa attggtatca gcagaaacca 120 gggaaagccc ctaacctcct
gatctatgct gcgtccagtt tacatggtgg ggtcccatca 180 aggttcagtg
gcagcggctc tgggacagat ttcactctca ccatcagaac tcttcaacct 240
gaagattttg caacttacta ctgtcaacag agttccaata ccccattcac tttcggccct
300 gggaccgtag tggatttcag a 321 <210> SEQ ID NO 170
<211> LENGTH: 107 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 170 Asp Ile Gln Met
Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Ser
Ile Thr Ile Thr Cys Arg Ala Ser Leu Ser Ile Asn Thr Phe 20 25 30
Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Asn Leu Leu Ile 35
40 45 Tyr Ala Ala Ser Ser Leu His Gly Gly Val Pro Ser Arg Phe Ser
Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Arg Thr
Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser
Ser Asn Thr Pro Phe 85 90 95 Thr Phe Gly Pro Gly Thr Val Val Asp
Phe Arg 100 105 <210> SEQ ID NO 171 <211> LENGTH: 18
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 171 ctgtccatta acaccttt 18 <210> SEQ ID
NO 172 <211> LENGTH: 6 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 172 Leu Ser Ile
Asn Thr Phe 1 5 <210> SEQ ID NO 173 <211> LENGTH: 9
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 173 gctgcgtcc 9 <210> SEQ ID NO 174
<211> LENGTH: 3 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 174 Ala Ala Ser 1
<210> SEQ ID NO 175 <211> LENGTH: 27 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 175
caacagagtt ccaatacccc attcact 27 <210> SEQ ID NO 176
<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 176 Gln Gln Ser Ser
Asn Thr Pro Phe Thr 1 5 <210> SEQ ID NO 177 <211>
LENGTH: 363 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 177 gaggtgcagc tggtggagtc
tgggggagga gtggtacggc cgggggggtc cctgagactc 60 tcctgtgcag
cctctggatt cacttttgat gactatggca tgagttgggt ccgccaagtt 120
ccagggaagg ggctggagtg ggtctcaggt attagttgga atgatggtaa gacagtttat
180 gcagagtctg tgaagggccg attcatcatc tccagagaca acgccaagaa
ctccctgtat 240 ctggaaatga atagtctgag agccgaggac acggccttat
attactgtgc gagagattgg 300 cagtacttga tagagcggta ctttgactac
tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210> SEQ ID NO
178 <211> LENGTH: 121 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 178 Glu Val Gln
Leu Val Glu Ser Gly Gly Gly Val Val Arg Pro Gly Gly 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25
30 Gly Met Ser Trp Val Arg Gln Val Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ser Gly Ile Ser Trp Asn Asp Gly Lys Thr Val Tyr Ala Glu
Ser Val 50 55 60 Lys Gly Arg Phe Ile Ile Ser Arg Asp Asn Ala Lys
Asn Ser Leu Tyr 65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp
Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Trp Gln Tyr Leu Ile
Glu Arg Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr
Val Ser Ser 115 120 <210> SEQ ID NO 179 <211> LENGTH:
24 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 179 ggattcactt ttgatgacta tggc 24 <210>
SEQ ID NO 180 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 180
Gly Phe Thr Phe Asp Asp Tyr Gly 1 5 <210> SEQ ID NO 181
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 181 attagttgga
atgatggtaa gaca 24 <210> SEQ ID NO 182 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 182 Ile Ser Trp Asn Asp Gly Lys Thr 1 5
<210> SEQ ID NO 183 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 183
gcgagagatt ggcagtactt gatagagcgg tactttgact ac 42 <210> SEQ
ID NO 184 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 184 Ala Arg Asp
Trp Gln Tyr Leu Ile Glu Arg Tyr Phe Asp Tyr 1 5 10 <210> SEQ
ID NO 185 <211> LENGTH: 324 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 185 gaaatagttt
tgacacagag tcccggcaca ctgtcactct ctcccgggga aagagccacc 60
ttgtcatgta gagcaagtca gtcagtctct agctcttatc tcgcctggta ccagcagaag
120 ccgggacagg cccctagact gctgatctac ggggcaagtt ccagggccac
cggaatcccc 180 gaccggttca gtggaagcgg aagcggaacc gattttactt
tgacgatttc tagactggag 240 ccagaggatt tcgccgttta ctattgtcaa
cagtacggaa gcagcccgtg gacgtttggc 300 cagggcacga aggtagaaat caag 324
<210> SEQ ID NO 186 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 186
Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5
10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser
Ser 20 25 30 Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro
Arg Leu Leu 35 40 45 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile
Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr
Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr
Tyr Cys Gln Gln Tyr Gly Ser Ser Pro 85 90 95 Trp Thr Phe Gly Gln
Gly Thr Lys Val Glu Ile Lys 100 105 <210> SEQ ID NO 187
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 187 cagtcagtct
ctagctctta t 21 <210> SEQ ID NO 188 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 188 Gln Ser Val Ser Ser Ser Tyr 1 5
<210> SEQ ID NO 189 <211> LENGTH: 9 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 189
ggggcaagt 9 <210> SEQ ID NO 190 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 190 Gly Ala Ser 1 <210> SEQ ID NO 191
<211> LENGTH: 27 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 191 caacagtacg
gaagcagccc gtggacg 27 <210> SEQ ID NO 192 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 192 Gln Gln Tyr Gly Ser Ser Pro Trp Thr 1 5
<210> SEQ ID NO 193 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 193
gaggtgcagc tggtggagtc tgggggaggt gtggtacggc ctggggggtc cctgagactc
60 tcctgtacag cctctggatt cacctttgat gattatggca tgagctgggt
ccgccaagct 120 ccagggaagg ggctggagtg gatctctggt attggttgga
ctggtggtcg gtcaagttat 180 gcagactctg tgaggggccg attcaccatc
tccagagaca acgccaagaa ttccctgtat 240 ctgcaaatga acagtctggg
agccgaggac acggccttgt attattgtgc aagagatcgg 300 cagtggctgg
tgcagtggta ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 194 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 194
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Arg Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Gly Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Ile 35 40 45 Ser Gly Ile Gly Trp Thr Gly Gly Arg Ser Ser
Tyr Ala Asp Ser Val 50 55 60 Arg Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Gly
Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Arg Gln
Trp Leu Val Gln Trp Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 195
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 195 ggattcacct
ttgatgatta tggc 24 <210> SEQ ID NO 196 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 196 Gly Phe Thr Phe Asp Asp Tyr Gly 1 5
<210> SEQ ID NO 197 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 197
attggttgga ctggtggtcg gtca 24 <210> SEQ ID NO 198 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 198 Ile Gly Trp Thr Gly Gly Arg Ser
1 5 <210> SEQ ID NO 199 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 199 gcaagagatc ggcagtggct ggtgcagtgg tactttgact ac 42
<210> SEQ ID NO 200 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 200
Ala Arg Asp Arg Gln Trp Leu Val Gln Trp Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 201 <211> LENGTH: 324 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 201
gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc
60 atcacttgcc gggcaagtca gagcattagc agctatttaa attggtatca
gcagaaacca 120 gggaaagccc ctaagctcct gatctatgct gcatccagtt
tgcaaagtgg ggtcccgtca 180 aggttcagtg gcagtggatc tgggacagat
ttcactctca ccatcagcag tctgcaacct 240 gaagattttg caacttacta
ctgtcaacag agttacagta cccctccgat caccttcggc 300 caagggacac
gactggagat taaa 324 <210> SEQ ID NO 202 <211> LENGTH:
108 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 202 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg
Ala Ser Gln Ser Ile Ser Ser Tyr 20 25 30 Leu Asn Trp Tyr Gln Gln
Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Ala Ala Ser
Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly
Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80
Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85
90 95 Ile Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys 100 105
<210> SEQ ID NO 203 <211> LENGTH: 18 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 203
cagagcatta gcagctat 18 <210> SEQ ID NO 204 <211>
LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 204 Gln Ser Ile Ser Ser Tyr 1 5
<210> SEQ ID NO 205 <211> LENGTH: 9 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 205
gctgcatcc 9 <210> SEQ ID NO 206 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 206 Ala Ala Ser 1 <210> SEQ ID NO 207
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 207 caacagagtt
acagtacccc tccgatcacc 30 <210> SEQ ID NO 208 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 208 Gln Gln Ser Tyr Ser Thr Pro Pro
Ile Thr 1 5 10 <210> SEQ ID NO 209 <211> LENGTH: 363
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 209 gaggtgcagc tggtggagtc tgggggaaga
gtggtacggc cgggggggtc cctgagactc 60 tcctgtgcag cctctggatt
cacttttgat gactatggca tgagttgggt ccgccaactt 120 ccagggaagg
gcctggagtg ggtcgcaggt attagttgga atgatggtaa gacagtttat 180
gcagagtctg tgaagggccg attcatcatc tccagagaca acgccaagaa ctccctgcat
240 ctggagatga acagtctgag agccgaggac acggccttat attactgtgc
gcgagattgg 300 caatacttaa tagatcgtta ctttgacttc tggggtcagg
gaaccctggt caccgtctcc 360 tca 363 <210> SEQ ID NO 210
<211> LENGTH: 121 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 210 Glu Val Gln Leu
Val Glu Ser Gly Gly Arg Val Val Arg Pro Gly Gly 1 5 10 15 Ser Leu
Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30
Gly Met Ser Trp Val Arg Gln Leu Pro Gly Lys Gly Leu Glu Trp Val 35
40 45 Ala Gly Ile Ser Trp Asn Asp Gly Lys Thr Val Tyr Ala Glu Ser
Val 50 55 60 Lys Gly Arg Phe Ile Ile Ser Arg Asp Asn Ala Lys Asn
Ser Leu His 65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp Thr
Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Trp Gln Tyr Leu Ile Asp
Arg Tyr Phe Asp Phe Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 211 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 211 ggattcactt ttgatgacta tggc 24 <210>
SEQ ID NO 212 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 212
Gly Phe Thr Phe Asp Asp Tyr Gly 1 5 <210> SEQ ID NO 213
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 213 attagttgga
atgatggtaa gaca 24 <210> SEQ ID NO 214 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 214 Ile Ser Trp Asn Asp Gly Lys Thr 1 5
<210> SEQ ID NO 215 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 215
gcgcgagatt ggcaatactt aatagatcgt tactttgact tc 42 <210> SEQ
ID NO 216 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 216 Ala Arg Asp
Trp Gln Tyr Leu Ile Asp Arg Tyr Phe Asp Phe 1 5 10 <210> SEQ
ID NO 217 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 217 gaagtgcagc
tggtggagtc tgggggaggc ttggtgcagc ctggcgggtc cctgagactc 60
tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct
120 ccagggaagg gcctggagtg ggtctcaggt attggttgga gtagtggtag
cataggctat 180 gcggactctg tgaagggccg attcaccatc tccagagaca
acgccaagaa ctccttgtat 240 ctgcaaatgg acagtctgag acctgaggac
tcagccttat attactgtgc aaaagcctat 300 acatttatga ttaccctcta
ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 218 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 218
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Gly Trp Ser Ser Gly Ser Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asp Ser Leu Arg
Pro Glu Asp Ser Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Ala Tyr Thr
Phe Met Ile Thr Leu Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 219
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 219 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 220 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 220 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 221 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 221
attggttgga gtagtggtag cata 24 <210> SEQ ID NO 222 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 222 Ile Gly Trp Ser Ser Gly Ser Ile
1 5 <210> SEQ ID NO 223 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 223 gcaaaagcct atacatttat gattaccctc tactttgact ac 42
<210> SEQ ID NO 224 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 224
Ala Lys Ala Tyr Thr Phe Met Ile Thr Leu Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 225 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 225
gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgat gattatgaca tgcactgggt
ccggcaagct 120 ccagggaagg gcctggagtg ggtgtcaggg agtggttgga
ataggggtag tttaggctat 180 gcggattctg tgaagggccg attcaccatc
tccagagaca acgccaagaa gtccctgtat 240 ctgcaaatga acagtgtgag
agttgaggac acggccttgt attactgtgc aaaaggcttt 300 gtagtggtat
cagctgctta ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 226 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 226
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Asp Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ser Gly Trp Asn Arg Gly Ser Leu Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Lys Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Val Arg
Val Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Gly Phe Val
Val Val Ser Ala Ala Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 227
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 227 ggattcacct
ttgatgatta tgac 24 <210> SEQ ID NO 228 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 228 Gly Phe Thr Phe Asp Asp Tyr Asp 1 5
<210> SEQ ID NO 229 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 229
agtggttgga ataggggtag ttta 24 <210> SEQ ID NO 230 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 230 Ser Gly Trp Asn Arg Gly Ser Leu
1 5 <210> SEQ ID NO 231 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 231 gcaaaaggct ttgtagtggt atcagctgct tactttgact ac 42
<210> SEQ ID NO 232 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 232
Ala Lys Gly Phe Val Val Val Ser Ala Ala Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 233 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 233
caggtgcagc tggtgcagtc tggggctgag gtgaagaggc ctgggtcctc ggtgaaggtc
60 tcctgcaagg tatctggagt caccttcagg aattttgcta tcatctgggt
gcgacaggcc 120 cctggacaag ggcttgagtg gatgggagga atcatccctt
tctttagtgc agcaaattac 180 gcacagagct tccagggcag agtcacgatt
accccggacg aatccacgag cacagccttc 240 atggagctgg ccagtctgag
atctgaggac acggccgttt attattgtgc gagagagggg 300 gaacgtggac
acacctatgg gtttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 234 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 234
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Arg Pro Gly Ser 1 5
10 15 Ser Val Lys Val Ser Cys Lys Val Ser Gly Val Thr Phe Arg Asn
Phe 20 25 30 Ala Ile Ile Trp Val Arg Gln Ala Pro Gly Gln Gly Leu
Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Phe Phe Ser Ala Ala Asn
Tyr Ala Gln Ser Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Pro Asp
Glu Ser Thr Ser Thr Ala Phe 65 70 75 80 Met Glu Leu Ala Ser Leu Arg
Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Glu Gly Glu
Arg Gly His Thr Tyr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 235
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 235 ggagtcacct
tcaggaattt tgct 24 <210> SEQ ID NO 236 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 236 Gly Val Thr Phe Arg Asn Phe Ala 1 5
<210> SEQ ID NO 237 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 237
atcatccctt tctttagtgc agca 24 <210> SEQ ID NO 238 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 238 Ile Ile Pro Phe Phe Ser Ala Ala
1 5 <210> SEQ ID NO 239 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 239 gcgagagagg gggaacgtgg acacacctat gggtttgact ac 42
<210> SEQ ID NO 240 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 240
Ala Arg Glu Gly Glu Arg Gly His Thr Tyr Gly Phe Asp Tyr 1 5 10
<210> SEQ ID NO 241 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 241
gaagtgcagc tggtggagtc tgggggaggc ttggtacagt ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt
ccgacaacct 120 ccagggaagg gcctggaatg ggtctcaggt attaactgga
atagaggtag gacaggctat 180 gcggactctg tgaagggccg attcaccatc
tccagagaca acgccaagaa ctccctgtat 240 ctgcaaatga acgatctgag
agttgaggat acggccttgt attactgtgc aaaagccgaa 300 cagtggctgg
acgagggata ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 242 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 242
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Ser Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Pro Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Asn Trp Asn Arg Gly Arg Thr Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Asp Leu Arg
Val Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Ala Glu Gln
Trp Leu Asp Glu Gly Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 243
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 243 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 244 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 244 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 245 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 245
attaactgga atagaggtag gaca 24 <210> SEQ ID NO 246 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 246 Ile Asn Trp Asn Arg Gly Arg Thr
1 5 <210> SEQ ID NO 247 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 247 gcaaaagccg aacagtggct ggacgaggga tactttgact ac 42
<210> SEQ ID NO 248 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 248
Ala Lys Ala Glu Gln Trp Leu Asp Glu Gly Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 249 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 249
gaggtgcagc tggtggagtc tgggggaggc ttggtgcagc ggggggggtc cctgagactc
60 tcctgtgcag cctctggatt cagctttagc agctatgcca tgaactgggt
ccgccaggct 120 ccagggaagg ggctggagtg ggtctcaact attagtgata
gtggtggtag tacatactac 180 gcagactccg tgaagggccg gttcaccatt
tccagagaca attccaagaa cacgctgtct 240 ctgcaaatga acagcctgag
agccgaggac acggccgtat attactgtgc gaaagatcag 300 ggtgggagtt
acccctacta ctttcactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 250 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 250
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Arg Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Ser Phe Ser Ser
Tyr 20 25 30 Ala Met Asn Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Thr Ile Ser Asp Ser Gly Gly Ser Thr Tyr
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ser Lys Asn Thr Leu Ser 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Gln Gly
Gly Ser Tyr Pro Tyr Tyr Phe His Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 251
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 251 ggattcagct
ttagcagcta tgcc 24 <210> SEQ ID NO 252 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 252 Gly Phe Ser Phe Ser Ser Tyr Ala 1 5
<210> SEQ ID NO 253 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 253
attagtgata gtggtggtag taca 24 <210> SEQ ID NO 254 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 254 Ile Ser Asp Ser Gly Gly Ser Thr
1 5 <210> SEQ ID NO 255 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 255 gcgaaagatc agggtgggag ttacccctac tactttcact ac 42
<210> SEQ ID NO 256 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 256
Ala Lys Asp Gln Gly Gly Ser Tyr Pro Tyr Tyr Phe His Tyr 1 5 10
<210> SEQ ID NO 257 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 257
gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgag gattatgcca tgcactgggt
ccggcaagct 120 ccagggaagg gcctggagtg ggtctcaggt attggttgga
gtaatgtaaa gataggctat 180 gcggactctg tgaagggccg attcaccatc
tccagagaca atgtcaggaa ctccctatat 240 ctgcaaatga acagtctgag
aactgaggac acggccttct attactgtgt aaaagcctat 300 acatctatgc
ttaccctcta ctttgactat tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 258 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 258
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Glu Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Gly Trp Ser Asn Val Lys Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Val Arg Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Thr Glu Asp Thr Ala Phe Tyr Tyr Cys 85 90 95 Val Lys Ala Tyr Thr
Ser Met Leu Thr Leu Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 259
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 259 ggattcacct
ttgaggatta tgcc 24 <210> SEQ ID NO 260 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 260 Gly Phe Thr Phe Glu Asp Tyr Ala 1 5
<210> SEQ ID NO 261 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 261
attggttgga gtaatgtaaa gata 24 <210> SEQ ID NO 262 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 262 Ile Gly Trp Ser Asn Val Lys Ile
1 5 <210> SEQ ID NO 263 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 263 gtaaaagcct atacatctat gcttaccctc tactttgact at 42
<210> SEQ ID NO 264 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 264
Val Lys Ala Tyr Thr Ser Met Leu Thr Leu Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 265 <211> LENGTH: 354 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 265
caggtgcagc tggtgcagtc tggggctgag gtgaagaggc ctggggcctc agtgaaggtt
60 tcctgcaagg catctggata caccttcacc agcttctata tgtactgggt
gcgacaggcc 120 cctggacaag ggcttgagtg gatgggaata atcaacccta
gtgatggtag cacaagcaac 180 gcacagaagt tccagggcag agtcaccatg
accagggaca cgtccacgag tacagtctac 240 atggagctga gcagcctgag
atctgaggac acggccgtgt attactgtgc gagacgggtg 300 gctggggata
tttttgatat ctggggccaa gggacaatgg tcaccgtctc ttca 354 <210>
SEQ ID NO 266 <211> LENGTH: 118 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 266
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Arg Pro Gly Ala 1 5
10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser
Phe 20 25 30 Tyr Met Tyr Trp Val Arg Gln Ala Pro Gly Gln Gly Leu
Glu Trp Met 35 40 45 Gly Ile Ile Asn Pro Ser Asp Gly Ser Thr Ser
Asn Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Arg Asp
Thr Ser Thr Ser Thr Val Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg
Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Arg Val Ala
Gly Asp Ile Phe Asp Ile Trp Gly Gln Gly Thr 100 105 110 Met Val Thr
Val Ser Ser 115 <210> SEQ ID NO 267 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 267 ggatacacct tcaccagctt ctat 24 <210>
SEQ ID NO 268 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 268
Gly Tyr Thr Phe Thr Ser Phe Tyr 1 5 <210> SEQ ID NO 269
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 269 atcaacccta
gtgatggtag caca 24 <210> SEQ ID NO 270 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 270 Ile Asn Pro Ser Asp Gly Ser Thr 1 5
<210> SEQ ID NO 271 <211> LENGTH: 33 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 271
gcgagacggg tggctgggga tatttttgat atc 33 <210> SEQ ID NO 272
<211> LENGTH: 11 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 272 Ala Arg Arg Val
Ala Gly Asp Ile Phe Asp Ile 1 5 10 <210> SEQ ID NO 273
<211> LENGTH: 357 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 273 caggtgcagc
tgcaggagtc gggcccagga ctggtgaagc cttcggagac cctgtccctc 60
acctgcactg tctctggtgg ctccatcagt agttaccact ggaactggat ccggcagagt
120 ccagggaagg gactggaatg gattggatat atctattata ttgggagcac
cgactataat 180 ccctccctcg agagtcgagt caccatatca gtagacacgt
ccaagaacca gttctccctg 240 aagctgagtt ctgtgaccgc tgcggacacg
gccgtgtatt actgtgcgag agtccccgtg 300 ggagctacag gggcttctga
tgtctggggc caagggacaa tggtcaccgt ctcttca 357 <210> SEQ ID NO
274 <211> LENGTH: 119 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 274 Gln Val Gln
Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr
Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr 20 25
30 His Trp Asn Trp Ile Arg Gln Ser Pro Gly Lys Gly Leu Glu Trp Ile
35 40 45 Gly Tyr Ile Tyr Tyr Ile Gly Ser Thr Asp Tyr Asn Pro Ser
Leu Glu 50 55 60 Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn
Gln Phe Ser Leu 65 70 75 80 Lys Leu Ser Ser Val Thr Ala Ala Asp Thr
Ala Val Tyr Tyr Cys Ala 85 90 95 Arg Val Pro Val Gly Ala Thr Gly
Ala Ser Asp Val Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser
Ser 115 <210> SEQ ID NO 275 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 275 ggtggctcca tcagtagtta ccac 24 <210>
SEQ ID NO 276 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 276
Gly Gly Ser Ile Ser Ser Tyr His 1 5 <210> SEQ ID NO 277
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 277 atctattata
ttgggagcac c 21 <210> SEQ ID NO 278 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 278 Ile Tyr Tyr Ile Gly Ser Thr 1 5
<210> SEQ ID NO 279 <211> LENGTH: 39 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 279
gcgagagtcc ccgtgggagc tacaggggct tctgatgtc 39 <210> SEQ ID NO
280 <211> LENGTH: 13 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 280 Ala Arg Val
Pro Val Gly Ala Thr Gly Ala Ser Asp Val 1 5 10 <210> SEQ ID
NO 281 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 281 gaggtgcagc
tggtggagtc tgggggaagt gtggttcgac ctggggggtc cctgagactc 60
tcctgtgtag tctctggatt cacctttgag gattatggtt tgagctgggt ccgccaaatt
120 ccagggaaag gactggagtg ggtctctggt attagttgga ctggtggtaa
cacaggttat 180 gcagactctg tgaagggccg cttcaccatc tccagagaca
acgccaagaa ctccctgtat 240 ctgcaaatga acagtctgag agccgaagac
acggccctgt atcactgtac gagagatcga 300 cagtggctga tgcagtggta
ttttgactat tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 282 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 282
Glu Val Gln Leu Val Glu Ser Gly Gly Ser Val Val Arg Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Val Val Ser Gly Phe Thr Phe Glu Asp
Tyr 20 25 30 Gly Leu Ser Trp Val Arg Gln Ile Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Ser Trp Thr Gly Gly Asn Thr Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Leu Tyr His Cys 85 90 95 Thr Arg Asp Arg Gln
Trp Leu Met Gln Trp Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 283
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 283 ggattcacct
ttgaggatta tggt 24 <210> SEQ ID NO 284 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 284 Gly Phe Thr Phe Glu Asp Tyr Gly 1 5
<210> SEQ ID NO 285 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 285
attagttgga ctggtggtaa caca 24 <210> SEQ ID NO 286 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 286 Ile Ser Trp Thr Gly Gly Asn Thr
1 5 <210> SEQ ID NO 287 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 287 acgagagatc gacagtggct gatgcagtgg tattttgact at 42
<210> SEQ ID NO 288 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 288
Thr Arg Asp Arg Gln Trp Leu Met Gln Trp Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 289 <211> LENGTH: 357 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 289
caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc
60 tcctgttcag cctctggatt caccttcagt gcctatgcca tgcactgggt
ccgccaggct 120 ccaggcaagg ggctggaatg ggtggcagct atctcatatg
gtggaagtga taaatactat 180 gcagactccg tgaagggccg attcaccatc
tccagagaca attccaagaa cacgctatat 240 ctgcaaatga acagcctgag
aactgacgac acggctgtgt attactgtgc gaaatccgct 300 cactggaact
tcttctttga ctactggggc cagggaaccc tggtcactgt ctcctca 357 <210>
SEQ ID NO 290 <211> LENGTH: 119 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 290
Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ser Ala Ser Gly Phe Thr Phe Ser Ala
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ala Ala Ile Ser Tyr Gly Gly Ser Asp Lys Tyr
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Thr Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Ala His
Trp Asn Phe Phe Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val
Thr Val Ser Ser 115 <210> SEQ ID NO 291 <211> LENGTH:
24 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 291 ggattcacct tcagtgccta tgcc 24 <210>
SEQ ID NO 292 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 292
Gly Phe Thr Phe Ser Ala Tyr Ala 1 5 <210> SEQ ID NO 293
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 293 atctcatatg
gtggaagtga taaa 24 <210> SEQ ID NO 294 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 294 Ile Ser Tyr Gly Gly Ser Asp Lys 1 5
<210> SEQ ID NO 295 <211> LENGTH: 36 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 295
gcgaaatccg ctcactggaa cttcttcttt gactac 36 <210> SEQ ID NO
296 <211> LENGTH: 12 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 296 Ala Lys Ser
Ala His Trp Asn Phe Phe Phe Asp Tyr 1 5 10 <210> SEQ ID NO
297 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 297 gaagtgcagc
tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc 60
tcctgtgtag cctctggatt cgcccttcat gattatgcca tgcactgggt ccggcaagtt
120 ccagggaagg gcctggagtg ggtctcaagt attagttgga atagtggtgt
cataggctat 180 gcggactctc tgaagggccg cttcaccatc tccagagaca
acgccaagaa ctccctgtat 240 ctgcaaatga acagtctgag agcagaggac
acggccttat actactgtgc aaaaggtagt 300 gggagctact acgtcagttg
gttcgacccc tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 298 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 298
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Phe Ala Leu His Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Val Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Ser Ile Ser Trp Asn Ser Gly Val Ile Gly
Tyr Ala Asp Ser Leu 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Gly Ser Gly
Ser Tyr Tyr Val Ser Trp Phe Asp Pro Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 299
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 299 ggattcgccc
ttcatgatta tgcc 24 <210> SEQ ID NO 300 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 300 Gly Phe Ala Leu His Asp Tyr Ala 1 5
<210> SEQ ID NO 301 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 301
attagttgga atagtggtgt cata 24 <210> SEQ ID NO 302 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 302 Ile Ser Trp Asn Ser Gly Val Ile
1 5 <210> SEQ ID NO 303 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 303 gcaaaaggta gtgggagcta ctacgtcagt tggttcgacc cc 42
<210> SEQ ID NO 304 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 304
Ala Lys Gly Ser Gly Ser Tyr Tyr Val Ser Trp Phe Asp Pro 1 5 10
<210> SEQ ID NO 305 <211> LENGTH: 369 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 305
cagctgcagc tgcaggagtc gggcccagga ctggttcagc cttcggagac cctgtccctc
60 acctgcactg tctctggtga ctccatcagt agtactgctt accactggga
ctggatccgc 120 cagccccccg ggaagggact ggagtggatt gggaccatca
cttataatgg gaacacctac 180 ttcaacccgt ccctcaagag tcgagtcacc
atatccgttg acacgtccaa gaaccagttc 240 tccctgaagc tactctctat
gaccgccgca gaaacggctg ttttttactg tgcgcgacat 300 ctaggatata
acagtgactt ctttcccttt gacttctggg gccagggaac cctggtcact 360
gtctcctca 369 <210> SEQ ID NO 306 <211> LENGTH: 123
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 306 Gln Leu Gln Leu Gln Glu Ser Gly Pro Gly
Leu Val Gln Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Thr Val
Ser Gly Asp Ser Ile Ser Ser Thr 20 25 30 Ala Tyr His Trp Asp Trp
Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu 35 40 45 Trp Ile Gly Thr
Ile Thr Tyr Asn Gly Asn Thr Tyr Phe Asn Pro Ser 50 55 60 Leu Lys
Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe 65 70 75 80
Ser Leu Lys Leu Leu Ser Met Thr Ala Ala Glu Thr Ala Val Phe Tyr 85
90 95 Cys Ala Arg His Leu Gly Tyr Asn Ser Asp Phe Phe Pro Phe Asp
Phe 100 105 110 Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120
<210> SEQ ID NO 307 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 307
ggtgactcca tcagtagtac tgcttaccac 30 <210> SEQ ID NO 308
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 308 Gly Asp Ser Ile
Ser Ser Thr Ala Tyr His 1 5 10 <210> SEQ ID NO 309
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 309 atcacttata
atgggaacac c 21 <210> SEQ ID NO 310 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 310 Ile Thr Tyr Asn Gly Asn Thr 1 5
<210> SEQ ID NO 311 <211> LENGTH: 45 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 311
gcgcgacatc taggatataa cagtgacttc tttccctttg acttc 45 <210>
SEQ ID NO 312 <211> LENGTH: 15 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 312
Ala Arg His Leu Gly Tyr Asn Ser Asp Phe Phe Pro Phe Asp Phe 1 5 10
15 <210> SEQ ID NO 313 <211> LENGTH: 357 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 313 gaggtgcagc tggtggagtc tgggggaggc ctggtacggc
cgggggggtc cctgagactc 60 tcctgtgcag cctctggatt cacctttagc
acctatgcca tggcctgggt ccgccagact 120 ccagggaagg ggctggaggg
ggtctcagct attgggggta gtggtgatag tacctattat 180 gtcgactccg
tgaagggccg gttcaccatc tccagggaca actccaagag cacgcttttt 240
ctgcaaatga atagcctgag agccgaggac acggccgttt attactgtgt gaaagtccgg
300 aattacgacg gttcttttga tatctggggc caagggacaa tggtcaccgt ctcttca
357 <210> SEQ ID NO 314 <211> LENGTH: 119 <212>
TYPE: PRT <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 314 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Arg
Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe
Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ala Trp Val Arg Gln Thr Pro
Gly Lys Gly Leu Glu Gly Val 35 40 45 Ser Ala Ile Gly Gly Ser Gly
Asp Ser Thr Tyr Tyr Val Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr
Ile Ser Arg Asp Asn Ser Lys Ser Thr Leu Phe 65 70 75 80 Leu Gln Met
Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val
Lys Val Arg Asn Tyr Asp Gly Ser Phe Asp Ile Trp Gly Gln Gly 100 105
110 Thr Met Val Thr Val Ser Ser 115 <210> SEQ ID NO 315
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 315 ggattcacct
ttagcaccta tgcc 24 <210> SEQ ID NO 316 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 316 Gly Phe Thr Phe Ser Thr Tyr Ala 1 5
<210> SEQ ID NO 317 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 317
attgggggta gtggtgatag tacc 24 <210> SEQ ID NO 318 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 318 Ile Gly Gly Ser Gly Asp Ser Thr
1 5 <210> SEQ ID NO 319 <211> LENGTH: 36 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 319 gtgaaagtcc ggaattacga cggttctttt gatatc 36
<210> SEQ ID NO 320 <211> LENGTH: 12 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 320
Val Lys Val Arg Asn Tyr Asp Gly Ser Phe Asp Ile 1 5 10 <210>
SEQ ID NO 321 <211> LENGTH: 174 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-1-MMH <220> FEATURE:
<223> OTHER INFORMATION: 1-146: aa 25-170 of NP_005009.2 with
C93S <220> FEATURE: <223> OTHER INFORMATION: 147-174:
myc-myc-hexahistidine <400> SEQUENCE: 321 Leu Asp Ser Pro Asp
Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Val
Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser
Asn Thr Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40
45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln
50 55 60 Pro Gly Gln Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn
Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn
Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro
Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val
Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro
Ser Pro Arg Pro Ala Gly Gln Phe Gln Thr 130 135 140 Leu Val Glu Gln
Lys Leu Ile Ser Glu Glu Asp Leu Gly Gly Glu Gln 145 150 155 160 Lys
Leu Ile Ser Glu Glu Asp Leu His His His His His His 165 170
<210> SEQ ID NO 322 <211> LENGTH: 174 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: MfPD-1-MMH <220> FEATURE:
<223> OTHER INFORMATION: 1-146: M.fascicularis PD-1 with C93S
<220> FEATURE: <223> OTHER INFORMATION: 147-174:
myc-myc-hexahistidine <400> SEQUENCE: 322 Leu Glu Ser Pro Asp
Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Leu
Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser
Asn Ala Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40
45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln
50 55 60 Pro Gly Arg Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn
Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn
Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro
Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val
Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro
Ser Pro Arg Pro Ala Gly Gln Phe Gln Ala 130 135 140 Leu Val Glu Gln
Lys Leu Ile Ser Glu Glu Asp Leu Gly Gly Glu Gln 145 150 155 160 Lys
Leu Ile Ser Glu Glu Asp Leu His His His His His His 165 170
<210> SEQ ID NO 323 <211> LENGTH: 379 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-1-mFc <220> FEATURE:
<223> OTHER INFORMATION: 1-146: aa 25-170 of NP-005009.2 with
C93S <220> FEATURE: <223> OTHER INFORMATION: 147-379:
mFc: aa 98-330 of P01863 <400> SEQUENCE: 323 Leu Asp Ser Pro
Asp Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu
Val Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30
Ser Asn Thr Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35
40 45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser
Gln 50 55 60 Pro Gly Gln Asp Ser Arg Phe Arg Val Thr Gln Leu Pro
Asn Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg
Asn Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala
Pro Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg
Val Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser
Pro Ser Pro Arg Pro Ala Gly Gln Phe Gln Thr 130 135 140 Leu Val Glu
Pro Arg Gly Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys 145 150 155 160
Cys Pro Ala Pro Asn Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro 165
170 175 Pro Lys Ile Lys Asp Val Leu Met Ile Ser Leu Ser Pro Ile Val
Thr 180 185 190 Cys Val Val Val Asp Val Ser Glu Asp Asp Pro Asp Val
Gln Ile Ser 195 200 205 Trp Phe Val Asn Asn Val Glu Val His Thr Ala
Gln Thr Gln Thr His 210 215 220 Arg Glu Asp Tyr Asn Ser Thr Leu Arg
Val Val Ser Ala Leu Pro Ile 225 230 235 240 Gln His Gln Asp Trp Met
Ser Gly Lys Glu Phe Lys Cys Lys Val Asn 245 250 255 Asn Lys Asp Leu
Pro Ala Pro Ile Glu Arg Thr Ile Ser Lys Pro Lys 260 265 270 Gly Ser
Val Arg Ala Pro Gln Val Tyr Val Leu Pro Pro Pro Glu Glu 275 280 285
Glu Met Thr Lys Lys Gln Val Thr Leu Thr Cys Met Val Thr Asp Phe 290
295 300 Met Pro Glu Asp Ile Tyr Val Glu Trp Thr Asn Asn Gly Lys Thr
Glu 305 310 315 320 Leu Asn Tyr Lys Asn Thr Glu Pro Val Leu Asp Ser
Asp Gly Ser Tyr 325 330 335 Phe Met Tyr Ser Lys Leu Arg Val Glu Lys
Lys Asn Trp Val Glu Arg 340 345 350 Asn Ser Tyr Ser Cys Ser Val Val
His Glu Gly Leu His Asn His His 355 360 365 Thr Thr Lys Ser Phe Ser
Arg Thr Pro Gly Lys 370 375 <210> SEQ ID NO 324 <211>
LENGTH: 373 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
hPD-1-hFc <220> FEATURE: <223> OTHER INFORMATION:
1-146: aa 25-170 of NP_005009.2 with C93S <220> FEATURE:
<223> OTHER INFORMATION: 147-373: hFc: aa 104-330 of P01857
<400> SEQUENCE: 324 Leu Asp Ser Pro Asp Arg Pro Trp Asn Pro
Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Val Val Thr Glu Gly Asp
Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser Asn Thr Ser Glu Ser
Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40 45 Ser Asn Gln Thr
Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln 50 55 60 Pro Gly
Gln Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn Gly Arg 65 70 75 80
Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn Asp Ser Gly Thr 85
90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro Lys Ala Gln Ile Lys
Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val Thr Glu Arg Arg Ala
Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro Ser Pro Arg Pro Ala
Gly Gln Phe Gln Thr 130 135 140 Leu Val Asp Lys Thr His Thr Cys Pro
Pro Cys Pro Ala Pro Glu Leu 145 150 155 160 Leu Gly Gly Pro Ser Val
Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr 165 170 175 Leu Met Ile Ser
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val 180 185 190 Ser His
Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val 195 200 205
Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser 210
215 220 Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp
Leu 225 230 235 240 Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys
Ala Leu Pro Ala 245 250 255 Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys
Gly Gln Pro Arg Glu Pro 260 265 270 Gln Val Tyr Thr Leu Pro Pro Ser
Arg Asp Glu Leu Thr Lys Asn Gln 275 280 285 Val Ser Leu Thr Cys Leu
Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala 290 295 300 Val Glu Trp Glu
Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr 305 310 315 320 Pro
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu 325 330
335 Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser
340 345 350 Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser
Leu Ser 355 360 365 Leu Ser Pro Gly Lys 370 <210> SEQ ID NO
325 <211> LENGTH: 448 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: hPD-L1-hFc <220> FEATURE: <223>
OTHER INFORMATION: 1-221: aa 19-239 of NP_054862.1 <220>
FEATURE: <223> OTHER INFORMATION: 222-448: hFc: aa 104-330 of
P01857 <400> SEQUENCE: 325 Phe Thr Val Thr Val Pro Lys Asp
Leu Tyr Val Val Glu Tyr Gly Ser 1 5 10 15 Asn Met Thr Ile Glu Cys
Lys Phe Pro Val Glu Lys Gln Leu Asp Leu 20 25 30 Ala Ala Leu Ile
Val Tyr Trp Glu Met Glu Asp Lys Asn Ile Ile Gln 35 40 45 Phe Val
His Gly Glu Glu Asp Leu Lys Val Gln His Ser Ser Tyr Arg 50 55 60
Gln Arg Ala Arg Leu Leu Lys Asp Gln Leu Ser Leu Gly Asn Ala Ala 65
70 75 80 Leu Gln Ile Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr
Arg Cys 85 90 95 Met Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile
Thr Val Lys Val 100 105 110 Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg
Ile Leu Val Val Asp Pro 115 120 125 Val Thr Ser Glu His Glu Leu Thr
Cys Gln Ala Glu Gly Tyr Pro Lys 130 135 140 Ala Glu Val Ile Trp Thr
Ser Ser Asp His Gln Val Leu Ser Gly Lys 145 150 155 160 Thr Thr Thr
Thr Asn Ser Lys Arg Glu Glu Lys Leu Phe Asn Val Thr 165 170 175 Ser
Thr Leu Arg Ile Asn Thr Thr Thr Asn Glu Ile Phe Tyr Cys Thr 180 185
190 Phe Arg Arg Leu Asp Pro Glu Glu Asn His Thr Ala Glu Leu Val Ile
195 200 205 Pro Glu Leu Pro Leu Ala His Pro Pro Asn Glu Arg Thr Asp
Lys Thr 210 215 220 His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
Gly Gly Pro Ser 225 230 235 240 Val Phe Leu Phe Pro Pro Lys Pro Lys
Asp Thr Leu Met Ile Ser Arg 245 250 255 Thr Pro Glu Val Thr Cys Val
Val Val Asp Val Ser His Glu Asp Pro 260 265 270 Glu Val Lys Phe Asn
Trp Tyr Val Asp Gly Val Glu Val His Asn Ala 275 280 285 Lys Thr Lys
Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val 290 295 300 Ser
Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr 305 310
315 320 Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys
Thr 325 330 335 Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val
Tyr Thr Leu 340 345 350 Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln
Val Ser Leu Thr Cys 355 360 365 Leu Val Lys Gly Phe Tyr Pro Ser Asp
Ile Ala Val Glu Trp Glu Ser 370 375 380 Asn Gly Gln Pro Glu Asn Asn
Tyr Lys Thr Thr Pro Pro Val Leu Asp 385 390 395 400 Ser Asp Gly Ser
Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser 405 410 415 Arg Trp
Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala 420 425 430
Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435
440 445 <210> SEQ ID NO 326 <211> LENGTH: 454
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: hPD-L1-mFc
<220> FEATURE: <223> OTHER INFORMATION: 1-221: aa
19-239 of NP_054862.1 <220> FEATURE: <223> OTHER
INFORMATION: 222-454: mFc: aa 98-330 of P01863 <400>
SEQUENCE: 326 Phe Thr Val Thr Val Pro Lys Asp Leu Tyr Val Val Glu
Tyr Gly Ser 1 5 10 15 Asn Met Thr Ile Glu Cys Lys Phe Pro Val Glu
Lys Gln Leu Asp Leu 20 25 30 Ala Ala Leu Ile Val Tyr Trp Glu Met
Glu Asp Lys Asn Ile Ile Gln 35 40 45 Phe Val His Gly Glu Glu Asp
Leu Lys Val Gln His Ser Ser Tyr Arg 50 55 60 Gln Arg Ala Arg Leu
Leu Lys Asp Gln Leu Ser Leu Gly Asn Ala Ala 65 70 75 80 Leu Gln Ile
Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr Arg Cys 85 90 95 Met
Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile Thr Val Lys Val 100 105
110 Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg Ile Leu Val Val Asp Pro
115 120 125 Val Thr Ser Glu His Glu Leu Thr Cys Gln Ala Glu Gly Tyr
Pro Lys 130 135 140 Ala Glu Val Ile Trp Thr Ser Ser Asp His Gln Val
Leu Ser Gly Lys 145 150 155 160 Thr Thr Thr Thr Asn Ser Lys Arg Glu
Glu Lys Leu Phe Asn Val Thr 165 170 175 Ser Thr Leu Arg Ile Asn Thr
Thr Thr Asn Glu Ile Phe Tyr Cys Thr 180 185 190 Phe Arg Arg Leu Asp
Pro Glu Glu Asn His Thr Ala Glu Leu Val Ile 195 200 205 Pro Glu Leu
Pro Leu Ala His Pro Pro Asn Glu Arg Thr Glu Pro Arg 210 215 220 Gly
Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys Cys Pro Ala Pro Asn 225 230
235 240 Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Ile Lys
Asp 245 250 255 Val Leu Met Ile Ser Leu Ser Pro Ile Val Thr Cys Val
Val Val Asp 260 265 270 Val Ser Glu Asp Asp Pro Asp Val Gln Ile Ser
Trp Phe Val Asn Asn 275 280 285 Val Glu Val His Thr Ala Gln Thr Gln
Thr His Arg Glu Asp Tyr Asn 290 295 300 Ser Thr Leu Arg Val Val Ser
Ala Leu Pro Ile Gln His Gln Asp Trp 305 310 315 320 Met Ser Gly Lys
Glu Phe Lys Cys Lys Val Asn Asn Lys Asp Leu Pro 325 330 335 Ala Pro
Ile Glu Arg Thr Ile Ser Lys Pro Lys Gly Ser Val Arg Ala 340 345 350
Pro Gln Val Tyr Val Leu Pro Pro Pro Glu Glu Glu Met Thr Lys Lys 355
360 365 Gln Val Thr Leu Thr Cys Met Val Thr Asp Phe Met Pro Glu Asp
Ile 370 375 380 Tyr Val Glu Trp Thr Asn Asn Gly Lys Thr Glu Leu Asn
Tyr Lys Asn 385 390 395 400 Thr Glu Pro Val Leu Asp Ser Asp Gly Ser
Tyr Phe Met Tyr Ser Lys 405 410 415 Leu Arg Val Glu Lys Lys Asn Trp
Val Glu Arg Asn Ser Tyr Ser Cys 420 425 430 Ser Val Val His Glu Gly
Leu His Asn His His Thr Thr Lys Ser Phe 435 440 445 Ser Arg Thr Pro
Gly Lys 450 <210> SEQ ID NO 327 <211> LENGTH: 288
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: hPD-1
NP_005009.2 <400> SEQUENCE: 327 Met Gln Ile Pro Gln Ala Pro
Trp Pro Val Val Trp Ala Val Leu Gln 1 5 10 15 Leu Gly Trp Arg Pro
Gly Trp Phe Leu Asp Ser Pro Asp Arg Pro Trp 20 25 30 Asn Pro Pro
Thr Phe Ser Pro Ala Leu Leu Val Val Thr Glu Gly Asp 35 40 45 Asn
Ala Thr Phe Thr Cys Ser Phe Ser Asn Thr Ser Glu Ser Phe Val 50 55
60 Leu Asn Trp Tyr Arg Met Ser Pro Ser Asn Gln Thr Asp Lys Leu Ala
65 70 75 80 Ala Phe Pro Glu Asp Arg Ser Gln Pro Gly Gln Asp Cys Arg
Phe Arg 85 90 95 Val Thr Gln Leu Pro Asn Gly Arg Asp Phe His Met
Ser Val Val Arg 100 105 110 Ala Arg Arg Asn Asp Ser Gly Thr Tyr Leu
Cys Gly Ala Ile Ser Leu 115 120 125 Ala Pro Lys Ala Gln Ile Lys Glu
Ser Leu Arg Ala Glu Leu Arg Val 130 135 140 Thr Glu Arg Arg Ala Glu
Val Pro Thr Ala His Pro Ser Pro Ser Pro 145 150 155 160 Arg Pro Ala
Gly Gln Phe Gln Thr Leu Val Val Gly Val Val Gly Gly 165 170 175 Leu
Leu Gly Ser Leu Val Leu Leu Val Trp Val Leu Ala Val Ile Cys 180 185
190 Ser Arg Ala Ala Arg Gly Thr Ile Gly Ala Arg Arg Thr Gly Gln Pro
195 200 205 Leu Lys Glu Asp Pro Ser Ala Val Pro Val Phe Ser Val Asp
Tyr Gly 210 215 220 Glu Leu Asp Phe Gln Trp Arg Glu Lys Thr Pro Glu
Pro Pro Val Pro 225 230 235 240 Cys Val Pro Glu Gln Thr Glu Tyr Ala
Thr Ile Val Phe Pro Ser Gly 245 250 255 Met Gly Thr Ser Ser Pro Ala
Arg Arg Gly Ser Ala Asp Gly Pro Arg 260 265 270 Ser Ala Gln Pro Leu
Arg Pro Glu Asp Gly His Cys Ser Trp Pro Leu 275 280 285 <210>
SEQ ID NO 328 <211> LENGTH: 290 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-L1 NP_054862.1 <400>
SEQUENCE: 328 Met Arg Ile Phe Ala Val Phe Ile Phe Met Thr Tyr Trp
His Leu Leu 1 5 10 15 Asn Ala Phe Thr Val Thr Val Pro Lys Asp Leu
Tyr Val Val Glu Tyr 20 25 30 Gly Ser Asn Met Thr Ile Glu Cys Lys
Phe Pro Val Glu Lys Gln Leu 35 40 45 Asp Leu Ala Ala Leu Ile Val
Tyr Trp Glu Met Glu Asp Lys Asn Ile 50 55 60 Ile Gln Phe Val His
Gly Glu Glu Asp Leu Lys Val Gln His Ser Ser 65 70 75 80 Tyr Arg Gln
Arg Ala Arg Leu Leu Lys Asp Gln Leu Ser Leu Gly Asn 85 90 95 Ala
Ala Leu Gln Ile Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr 100 105
110 Arg Cys Met Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile Thr Val
115 120 125 Lys Val Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg Ile Leu
Val Val 130 135 140 Asp Pro Val Thr Ser Glu His Glu Leu Thr Cys Gln
Ala Glu Gly Tyr 145 150 155 160 Pro Lys Ala Glu Val Ile Trp Thr Ser
Ser Asp His Gln Val Leu Ser 165 170 175 Gly Lys Thr Thr Thr Thr Asn
Ser Lys Arg Glu Glu Lys Leu Phe Asn 180 185 190 Val Thr Ser Thr Leu
Arg Ile Asn Thr Thr Thr Asn Glu Ile Phe Tyr 195 200 205 Cys Thr Phe
Arg Arg Leu Asp Pro Glu Glu Asn His Thr Ala Glu Leu 210 215 220 Val
Ile Pro Glu Leu Pro Leu Ala His Pro Pro Asn Glu Arg Thr His 225 230
235 240 Leu Val Ile Leu Gly Ala Ile Leu Leu Cys Leu Gly Val Ala Leu
Thr 245 250 255 Phe Ile Phe Arg Leu Arg Lys Gly Arg Met Met Asp Val
Lys Lys Cys 260 265 270 Gly Ile Gln Asp Thr Asn Ser Lys Lys Gln Ser
Asp Thr His Leu Glu 275 280 285 Glu Thr 290 <210> SEQ ID NO
329 <211> LENGTH: 379 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: aa1-146: M.fascicularis PD-1 (with C93S change)
aa147-379: mFc tag (aa 98-330 of P01863) <400> SEQUENCE: 329
Leu Glu Ser Pro Asp Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5
10 15 Leu Leu Leu Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser
Phe 20 25 30 Ser Asn Ala Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg
Met Ser Pro 35 40 45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro
Glu Asp Arg Ser Gln 50 55 60 Pro Gly Arg Asp Ser Arg Phe Arg Val
Thr Gln Leu Pro Asn Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val
Arg Ala Arg Arg Asn Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala
Ile Ser Leu Ala Pro Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg
Ala Glu Leu Arg Val Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr
Ala His Pro Ser Pro Ser Pro Arg Pro Ala Gly Gln Phe Gln Ala 130 135
140 Leu Val Glu Pro Arg Gly Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys
145 150 155 160 Cys Pro Ala Pro Asn Leu Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro 165 170 175 Pro Lys Ile Lys Asp Val Leu Met Ile Ser Leu
Ser Pro Ile Val Thr 180 185 190 Cys Val Val Val Asp Val Ser Glu Asp
Asp Pro Asp Val Gln Ile Ser 195 200 205 Trp Phe Val Asn Asn Val Glu
Val His Thr Ala Gln Thr Gln Thr His 210 215 220 Arg Glu Asp Tyr Asn
Ser Thr Leu Arg Val Val Ser Ala Leu Pro Ile 225 230 235 240 Gln His
Gln Asp Trp Met Ser Gly Lys Glu Phe Lys Cys Lys Val Asn 245 250 255
Asn Lys Asp Leu Pro Ala Pro Ile Glu Arg Thr Ile Ser Lys Pro Lys 260
265 270 Gly Ser Val Arg Ala Pro Gln Val Tyr Val Leu Pro Pro Pro Glu
Glu 275 280 285 Glu Met Thr Lys Lys Gln Val Thr Leu Thr Cys Met Val
Thr Asp Phe 290 295 300 Met Pro Glu Asp Ile Tyr Val Glu Trp Thr Asn
Asn Gly Lys Thr Glu 305 310 315 320 Leu Asn Tyr Lys Asn Thr Glu Pro
Val Leu Asp Ser Asp Gly Ser Tyr 325 330 335 Phe Met Tyr Ser Lys Leu
Arg Val Glu Lys Lys Asn Trp Val Glu Arg 340 345 350 Asn Ser Tyr Ser
Cys Ser Val Val His Glu Gly Leu His Asn His His 355 360 365 Thr Thr
Lys Ser Phe Ser Arg Thr Pro Gly Lys 370 375 <210> SEQ ID NO
330 <211> LENGTH: 444 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: H4H7798N aa1-117: HCVR aa118-444: HC constant
<400> SEQUENCE: 330 Glu Val Gln Leu Leu Glu Ser Gly Gly Val
Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe Thr Phe Ser Asn Phe 20 25 30 Gly Met Thr Trp Val Arg
Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser
Gly Gly Gly Arg Asp Thr Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly
Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80
Leu Gln Met Asn Ser Leu Lys Gly Glu Asp Thr Ala Val Tyr Tyr Cys 85
90 95 Val Lys Trp Gly Asn Ile Tyr Phe Asp Tyr Trp Gly Gln Gly Thr
Leu 100 105 110 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val
Phe Pro Leu 115 120 125 Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr
Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro
Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly
Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr
Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly
Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn 195 200 205
Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro 210
215 220 Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu
Phe 225 230 235 240 Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg
Thr Pro Glu Val 245 250 255 Thr Cys Val Val Val Asp Val Ser Gln Glu
Asp Pro Glu Val Gln Phe 260 265 270 Asn Trp Tyr Val Asp Gly Val Glu
Val His Asn Ala Lys Thr Lys Pro 275 280 285 Arg Glu Glu Gln Phe Asn
Ser Thr Tyr Arg Val Val Ser Val Leu Thr 290 295 300 Val Leu His Gln
Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val 305 310 315 320 Ser
Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala 325 330
335 Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln
340 345 350 Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val
Lys Gly 355 360 365 Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser
Asn Gly Gln Pro 370 375 380 Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val
Leu Asp Ser Asp Gly Ser 385 390 395 400 Phe Phe Leu Tyr Ser Arg Leu
Thr Val Asp Lys Ser Arg Trp Gln Glu 405 410 415 Gly Asn Val Phe Ser
Cys Ser Val Met His Glu Ala Leu His Asn His 420 425 430 Tyr Thr Gln
Lys Ser Leu Ser Leu Ser Leu Gly Lys 435 440 <210> SEQ ID NO
331 <211> LENGTH: 214 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: H4H7798N aa1-107: LCVR aa108-214: LC constant
<400> SEQUENCE: 331 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Ser Ile Thr Ile Thr Cys Arg
Ala Ser Leu Ser Ile Asn Thr Phe 20 25 30 Leu Asn Trp Tyr Gln Gln
Lys Pro Gly Lys Ala Pro Asn Leu Leu Ile 35 40 45 Tyr Ala Ala Ser
Ser Leu His Gly Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly
Ser Gly Thr Asp Phe Thr Leu Thr Ile Arg Thr Leu Gln Pro 65 70 75 80
Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Ser Asn Thr Pro Phe 85
90 95 Thr Phe Gly Pro Gly Thr Val Val Asp Phe Arg Arg Thr Val Ala
Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu
Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe
Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val Asp Asn Ala
Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val Thr Glu Gln
Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr
Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys
Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205
Phe Asn Arg Gly Glu Cys 210 <210> SEQ ID NO 332 <211>
LENGTH: 449 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
H4H7795N2 aa1-122: HCVR aa123-449: HC constant <400>
SEQUENCE: 332 Glu Val Gln Val Val Glu Ser Gly Gly Gly Leu Val Glu
Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Lys Ala Ser Gly Phe
Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Thr Pro
Gly Lys Ala Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Trp Ser Gly
Asn Asn Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr
Ile Ser Lys Asp Asp Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met
Asn Ser Leu Arg Pro Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Thr
Lys Asp Ile Ser Ile Thr Gly Thr Leu Asp Ala Phe Asp Val Trp 100 105
110 Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro
115 120 125 Ser Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu
Ser Thr 130 135 140 Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro
Glu Pro Val Thr 145 150 155 160 Val Ser Trp Asn Ser Gly Ala Leu Thr
Ser Gly Val His Thr Phe Pro 165 170 175 Ala Val Leu Gln Ser Ser Gly
Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190 Val Pro Ser Ser Ser
Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp 195 200 205 His Lys Pro
Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr 210 215 220 Gly
Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro 225 230
235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile
Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
Gln Glu Asp 260 265 270 Pro Glu Val Gln Phe Asn Trp Tyr Val Asp Gly
Val Glu Val His Asn 275 280 285 Ala Lys Thr Lys Pro Arg Glu Glu Gln
Phe Asn Ser Thr Tyr Arg Val 290 295 300 Val Ser Val Leu Thr Val Leu
His Gln Asp Trp Leu Asn Gly Lys Glu 305 310 315 320 Tyr Lys Cys Lys
Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys 325 330 335 Thr Ile
Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 340 345 350
Leu Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr 355
360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp
Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
Pro Val Leu 385 390 395 400 Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser
Arg Leu Thr Val Asp Lys 405 410 415 Ser Arg Trp Gln Glu Gly Asn Val
Phe Ser Cys Ser Val Met His Glu 420 425 430 Ala Leu His Asn His Tyr
Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly 435 440 445 Lys <210>
SEQ ID NO 333 <211> LENGTH: 214 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H7795N2 aa1-107: LCVR aa108-214:
LC constant <400> SEQUENCE: 333 Asp Ile Gln Met Thr Gln Ser
Pro Ile Ser Val Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile
Thr Cys Arg Ala Ser Gln Gly Ile Ser Asn Trp 20 25 30 Leu Ala Trp
Tyr Gln Gln Lys Pro Gly Ile Ala Pro Lys Leu Leu Ile 35 40 45 Tyr
Ser Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Arg Gly 50 55
60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Gly Ser Leu Gln Pro
65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ala His Ser Phe
Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg
Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp
Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu
Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val
Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val
Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser
Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185
190 Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser
195 200 205 Phe Asn Arg Gly Glu Cys 210 <210> SEQ ID NO 334
<211> LENGTH: 446 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: H4H9008P aa1-119: HCVR aa120-446: HC constant
<400> SEQUENCE: 334 Glu Val Gln Leu Val Glu Ser Gly Gly Gly
Leu Val Arg Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ala Trp Val Arg
Gln Thr Pro Gly Lys Gly Leu Glu Gly Val 35 40 45 Ser Ala Ile Gly
Gly Ser Gly Asp Ser Thr Tyr Tyr Val Asp Ser Val 50 55 60 Lys Gly
Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Ser Thr Leu Phe 65 70 75 80
Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85
90 95 Val Lys Val Arg Asn Tyr Asp Gly Ser Phe Asp Ile Trp Gly Gln
Gly 100 105 110 Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro
Ser Val Phe 115 120 125 Pro Leu Ala Pro Cys Ser Arg Ser Thr Ser Glu
Ser Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp Tyr Phe Pro
Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly Ala Leu Thr
Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Ser Gly
Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185 190 Ser Ser
Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro 195 200 205
Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro 210
215 220 Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val
Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile
Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser
Gln Glu Asp Pro Glu Val 260 265 270 Gln Phe Asn Trp Tyr Val Asp Gly
Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln
Phe Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu
His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys
Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser 325 330
335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro
340 345 350 Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys
Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp
Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser
Arg Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Glu Gly Asn Val
Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr
Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys 435 440 445 <210>
SEQ ID NO 335 <211> LENGTH: 215 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H9008P aa1-108: LCVR aa109-215: LC
constant <400> SEQUENCE: 335 Glu Ile Val Leu Thr Gln Ser Pro
Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Ser 20 25 30 Tyr Leu Ala Trp
Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45 Ile Tyr
Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser 50 55 60
Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu 65
70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Gly Ser
Ser Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys
Arg Thr Val Ala 100 105 110 Ala Pro Ser Val Phe Ile Phe Pro Pro Ser
Asp Glu Gln Leu Lys Ser 115 120 125 Gly Thr Ala Ser Val Val Cys Leu
Leu Asn Asn Phe Tyr Pro Arg Glu 130 135 140 Ala Lys Val Gln Trp Lys
Val Asp Asn Ala Leu Gln Ser Gly Asn Ser 145 150 155 160 Gln Glu Ser
Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu 165 170 175 Ser
Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val 180 185
190 Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys
195 200 205 Ser Phe Asn Arg Gly Glu Cys 210 215 <210> SEQ ID
NO 336 <211> LENGTH: 448 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: H4H9048P2 aa1-121: HCVR aa122-448: HC constant
<400> SEQUENCE: 336 Gln Val Gln Leu Val Gln Ser Gly Ala Glu
Val Lys Arg Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Val
Ser Gly Val Thr Phe Arg Asn Phe 20 25 30 Ala Ile Ile Trp Val Arg
Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile
Pro Phe Phe Ser Ala Ala Asn Tyr Ala Gln Ser Phe 50 55 60 Gln Gly
Arg Val Thr Ile Thr Pro Asp Glu Ser Thr Ser Thr Ala Phe 65 70 75 80
Met Glu Leu Ala Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85
90 95 Ala Arg Glu Gly Glu Arg Gly His Thr Tyr Gly Phe Asp Tyr Trp
Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys
Gly Pro Ser 115 120 125 Val Phe Pro Leu Ala Pro Cys Ser Arg Ser Thr
Ser Glu Ser Thr Ala 130 135 140 Ala Leu Gly Cys Leu Val Lys Asp Tyr
Phe Pro Glu Pro Val Thr Val 145 150 155 160 Ser Trp Asn Ser Gly Ala
Leu Thr Ser Gly Val His Thr Phe Pro Ala 165 170 175 Val Leu Gln Ser
Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 180 185 190 Pro Ser
Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His 195 200 205
Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly 210
215 220 Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro
Ser 225 230 235 240 Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
Met Ile Ser Arg 245 250 255 Thr Pro Glu Val Thr Cys Val Val Val Asp
Val Ser Gln Glu Asp Pro 260 265 270 Glu Val Gln Phe Asn Trp Tyr Val
Asp Gly Val Glu Val His Asn Ala 275 280 285 Lys Thr Lys Pro Arg Glu
Glu Gln Phe Asn Ser Thr Tyr Arg Val Val 290 295 300 Ser Val Leu Thr
Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr 305 310 315 320 Lys
Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr 325 330
335 Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu
340 345 350 Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser Leu
Thr Cys 355 360 365 Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
Glu Trp Glu Ser 370 375 380 Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr
Thr Pro Pro Val Leu Asp 385 390 395 400 Ser Asp Gly Ser Phe Phe Leu
Tyr Ser Arg Leu Thr Val Asp Lys Ser 405 410 415 Arg Trp Gln Glu Gly
Asn Val Phe Ser Cys Ser Val Met His Glu Ala 420 425 430 Leu His Asn
His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys 435 440 445
<210> SEQ ID NO 337 <211> LENGTH: 215 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H9048P2 aa1-108: LCVR aa109-215:
LC constant <400> SEQUENCE: 337 Asp Ile Gln Met Thr Gln Ser
Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile
Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr 20 25 30 Leu Asn Trp
Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr
Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55
60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro
65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr
Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys
Arg Thr Val Ala 100 105 110 Ala Pro Ser Val Phe Ile Phe Pro Pro Ser
Asp Glu Gln Leu Lys Ser 115 120 125 Gly Thr Ala Ser Val Val Cys Leu
Leu Asn Asn Phe Tyr Pro Arg Glu 130 135 140 Ala Lys Val Gln Trp Lys
Val Asp Asn Ala Leu Gln Ser Gly Asn Ser 145 150 155 160 Gln Glu Ser
Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu 165 170 175 Ser
Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val 180 185
190 Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys
195 200 205 Ser Phe Asn Arg Gly Glu Cys 210 215
1 SEQUENCE LISTING <160> NUMBER OF SEQ ID NOS: 337
<210> SEQ ID NO 1 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 1
gaggtgcagc tgttggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc
60 tcctgttcag cctctggatt cacctttagc agctatacca tgaactgggt
ccgccaggct 120 ccagggaagg ggctggagtg ggtctcaggt attagtgata
ccggtggtaa cacatactac 180 acagactccg tgaagggccg gttcaccgtc
tccagagaca attccaagaa cacactgtct 240 ctgcaaatga acagcctgag
agccgaggac acggccgtat attactgtgc gaaagatcag 300 ggtggaagtt
acccctatta ctttcactac tggggccagg gatccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 2 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 2
Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ser Ala Ser Gly Phe Thr Phe Ser Ser
Tyr 20 25 30 Thr Met Asn Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Ser Asp Thr Gly Gly Asn Thr Tyr
Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Arg Asp
Asn Ser Lys Asn Thr Leu Ser 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Gln Gly
Gly Ser Tyr Pro Tyr Tyr Phe His Tyr Trp Gly 100 105 110 Gln Gly Ser
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 3 <211>
LENGTH: 24 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 3 ggattcacct ttagcagcta tacc 24
<210> SEQ ID NO 4 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 4
Gly Phe Thr Phe Ser Ser Tyr Thr 1 5 <210> SEQ ID NO 5
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 5 attagtgata
ccggtggtaa caca 24 <210> SEQ ID NO 6 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 6 Ile Ser Asp Thr Gly Gly Asn Thr 1 5
<210> SEQ ID NO 7 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 7
gcgaaagatc agggtggaag ttacccctat tactttcact ac 42 <210> SEQ
ID NO 8 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 8 Ala Lys Asp
Gln Gly Gly Ser Tyr Pro Tyr Tyr Phe His Tyr 1 5 10 <210> SEQ
ID NO 9 <211> LENGTH: 324 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 9 gacatccaga
tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60
atcacttgcc gggcaagtca gagcattagc agctatttaa tttggtatca gcagaaacca
120 gggacagccc ctaagttcct gatctatgct gcatccagtt tgcaaagtgg
ggtcccatca 180 aggttcagtg gctgtggatc tgggacagat ttcactctca
ccatcagcag tctgcaacct 240 gaagattttg caacttacta ctgtcaacag
agttacagta cccctccgat caccttcggc 300 caagggacac gactggagat taaa 324
<210> SEQ ID NO 10 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 10
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser
Tyr 20 25 30 Leu Ile Trp Tyr Gln Gln Lys Pro Gly Thr Ala Pro Lys
Phe Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Cys Gly Ser Gly Thr Asp Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln
Gly Thr Arg Leu Glu Ile Lys 100 105 <210> SEQ ID NO 11
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 11 cagagcatta gcagctat
18 <210> SEQ ID NO 12 <211> LENGTH: 6 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 12
Gln Ser Ile Ser Ser Tyr 1 5 <210> SEQ ID NO 13 <211>
LENGTH: 9 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 13 gctgcatcc 9 <210> SEQ ID
NO 14 <211> LENGTH: 3 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 14 Ala Ala Ser 1
<210> SEQ ID NO 15 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE:
15
caacagagtt acagtacccc tccgatcacc 30 <210> SEQ ID NO 16
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 16 Gln Gln Ser Tyr Ser
Thr Pro Pro Ile Thr 1 5 10 <210> SEQ ID NO 17 <211>
LENGTH: 366 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 17 gaggtgcagc tggtggagtc tggaggaggc
ttggtccagc ctggggggtc cctgagactc 60 tcctgtgcag cctctgggtt
caccgtcagt aacaactaca tgagctgggt ccgccaggct 120 ccagggaagg
ggctggagtg ggtctcagtt atttatagcg gtggtttcac atactacaca 180
gactccgtga agggccgatt caccatctcc agacacaatt ccaagaacac gctgtatctt
240 caaatgaaca gcctgagagc tgaggacacg gccgtgtatt actgtgcgag
gtattactat 300 gatactagtg attattggac cttctttgac tactggggcc
agggaaccct ggtcaccgtc 360 tcctca 366 <210> SEQ ID NO 18
<211> LENGTH: 122 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 18 Glu Val Gln Leu Val
Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg
Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Asn Asn 20 25 30 Tyr
Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40
45 Ser Val Ile Tyr Ser Gly Gly Phe Thr Tyr Tyr Thr Asp Ser Val Lys
50 55 60 Gly Arg Phe Thr Ile Ser Arg His Asn Ser Lys Asn Thr Leu
Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val
Tyr Tyr Cys Ala 85 90 95 Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp
Thr Phe Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 19 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 19 gggttcaccg tcagtaacaa ctac 24 <210>
SEQ ID NO 20 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 20
Gly Phe Thr Val Ser Asn Asn Tyr 1 5 <210> SEQ ID NO 21
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 21 atttatagcg
gtggtttcac a 21 <210> SEQ ID NO 22 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 22 Ile Tyr Ser Gly Gly Phe Thr 1 5
<210> SEQ ID NO 23 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 23
gcgaggtatt actatgatac tagtgattat tggaccttct ttgactac 48 <210>
SEQ ID NO 24 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 24
Ala Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp Thr Phe Phe Asp Tyr 1 5
10 15 <210> SEQ ID NO 25 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 25 gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga
aagagccacc 60 ctctcctgca gggccagtca gagtgttagc agcaacttag
cctggtacca gcagaaacct 120 ggccaggctc ccaggctcct catctatggt
gcatccacca gggccactgg tatcccagcc 180 aggttcagtg gcagtgggtc
tgggacagag ttcactctca ccatcagtag cctgcagtct 240 ggagattttg
cagtttatta ctgtcagcag tataataact ggccgctcac tttcggcgga 300
gggaccaagg tggagatcaa t 321 <210> SEQ ID NO 26 <211>
LENGTH: 107 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 26 Glu Ile Val Met Thr Gln Ser Pro
Ala Thr Leu Ser Val Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Asn 20 25 30 Leu Ala Trp Tyr
Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Gly
Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser 65
70 75 80 Gly Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn Asn Trp
Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Asn 100
105 <210> SEQ ID NO 27 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 27 cagagtgtta gcagcaac 18 <210> SEQ ID NO 28
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 28 Gln Ser Val Ser Ser
Asn 1 5 <210> SEQ ID NO 29 <211> LENGTH: 9 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 29 ggtgcatcc 9 <210> SEQ ID NO 30 <211>
LENGTH: 3 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 30 Gly Ala Ser 1
<210> SEQ ID NO 31 <211> LENGTH: 27 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 31
cagcagtata ataactggcc gctcact 27 <210> SEQ ID NO 32
<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 32 Gln Gln Tyr Asn Asn
Trp Pro Leu Thr 1 5 <210> SEQ ID NO 33 <211> LENGTH:
366 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 33 gaggtgcagc tggtggagtc tggaggaggc
ttggtccagc ctggggggtc cctgagactc 60 tcctgtgcag cctctgggtt
caccgtcagt aacaactaca tgagctgggt ccgccaggct 120 ccagggaagg
ggctggagtg ggtctcagtt atttatagcg gtggtttcac atactacaca 180
gactccgtga agggccgatt caccatctcc agacacaatt ccaagaacac gctgtatctt
240 caaatgaaca gcctgagagc tgaggacacg gccgtgtatt actgtgcgag
gtattactat 300 gatactagtg attattggac cttctttgac tactggggcc
agggaaccct ggtcaccgtc 360 tcctca 366 <210> SEQ ID NO 34
<211> LENGTH: 122 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 34 Glu Val Gln Leu Val
Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg
Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Asn Asn 20 25 30 Tyr
Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40
45 Ser Val Ile Tyr Ser Gly Gly Phe Thr Tyr Tyr Thr Asp Ser Val Lys
50 55 60 Gly Arg Phe Thr Ile Ser Arg His Asn Ser Lys Asn Thr Leu
Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val
Tyr Tyr Cys Ala 85 90 95 Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp
Thr Phe Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 35 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 35 gggttcaccg tcagtaacaa ctac 24 <210>
SEQ ID NO 36 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 36
Gly Phe Thr Val Ser Asn Asn Tyr 1 5 <210> SEQ ID NO 37
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 37 atttatagcg
gtggtttcac a 21 <210> SEQ ID NO 38 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 38 Ile Tyr Ser Gly Gly Phe Thr 1 5
<210> SEQ ID NO 39 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 39
gcgaggtatt actatgatac tagtgattat tggaccttct ttgactac 48 <210>
SEQ ID NO 40 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 40
Ala Arg Tyr Tyr Tyr Asp Thr Ser Asp Tyr Trp Thr Phe Phe Asp Tyr 1 5
10 15 <210> SEQ ID NO 41 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 41 gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga
aagagccacc 60 ctctcctgca gggccagtca gagtgttagc agcaacttag
cctggtacca gcagaaacct 120 ggccaggctc ccaggctcct catctatggt
gcatccacca gggccactgg tatcccagcc 180 aggttcagtg gcagtgggtc
tgggacagag ttcactctca ccatcagtag cctgcagtct 240 ggagattttg
cagtttatta ctgtcagcag tataataact ggccgctcac tttcggcgga 300
gggaccaagg tggagatcaa t 321 <210> SEQ ID NO 42 <211>
LENGTH: 107 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 42 Glu Ile Val Met Thr Gln Ser Pro
Ala Thr Leu Ser Val Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Asn 20 25 30 Leu Ala Trp Tyr
Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Gly
Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser 65
70 75 80 Gly Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn Asn Trp
Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Asn 100
105 <210> SEQ ID NO 43 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 43 cagagtgtta gcagcaac 18 <210> SEQ ID NO 44
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 44 Gln Ser Val Ser Ser
Asn 1 5 <210> SEQ ID NO 45 <211> LENGTH: 9 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 45 ggtgcatcc 9 <210> SEQ ID NO 46
<211> LENGTH: 3 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 46 Gly Ala Ser 1
<210> SEQ ID NO 47 <211> LENGTH: 27 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 47
cagcagtata ataactggcc gctcact 27 <210> SEQ ID NO 48
<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 48 Gln Gln Tyr Asn Asn
Trp Pro Leu Thr 1 5 <210> SEQ ID NO 49 <211> LENGTH:
357 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 49 caggtgcagc tggtggagtc tgggggaggc
gtggtccagt ctgggaggtc cctgagactc 60 tcctgtgcag cgtctggatt
caccttcagt agctatggca tgcactgggt ccgccaggct 120 ccaggcaagg
ggctggagtg ggtggcagtt atatggtatg atggaagtaa tatatactat 180
tcagactccg tgaagggccg attcaccatc tccagagcca attccaagaa cacgctgtat
240 ctgcaaatga acagcctgag agccgaggac acggctgttt attactgtgc
gagaccggga 300 cactggaact acttctttga atactggggc cagggaaccc
tggtcaccgt ctcctca 357 <210> SEQ ID NO 50 <211> LENGTH:
119 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 50 Gln Val Gln Leu Val Glu Ser Gly Gly Gly
Val Val Gln Ser Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Gly Met His Trp Val Arg
Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Trp
Tyr Asp Gly Ser Asn Ile Tyr Tyr Ser Asp Ser Val 50 55 60 Lys Gly
Arg Phe Thr Ile Ser Arg Ala Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80
Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85
90 95 Ala Arg Pro Gly His Trp Asn Tyr Phe Phe Glu Tyr Trp Gly Gln
Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> SEQ ID
NO 51 <211> LENGTH: 24 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 51 ggattcacct
tcagtagcta tggc 24 <210> SEQ ID NO 52 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 52 Gly Phe Thr Phe Ser Ser Tyr Gly 1 5
<210> SEQ ID NO 53 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 53
atatggtatg atggaagtaa tata 24 <210> SEQ ID NO 54 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 54 Ile Trp Tyr Asp Gly Ser Asn Ile
1 5 <210> SEQ ID NO 55 <211> LENGTH: 36 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 55 gcgagaccgg gacactggaa ctacttcttt gaatac 36 <210>
SEQ ID NO 56 <211> LENGTH: 12 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 56
Ala Arg Pro Gly His Trp Asn Tyr Phe Phe Glu Tyr 1 5 10 <210>
SEQ ID NO 57 <211> LENGTH: 324 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 57
gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc
60 atcacttgcc gggcaagtca gagcattaac aactatttaa attggtatca
gcagaaacca 120 gggaaagccc ctaagctcct gatctatact gcatccagtt
tgcaaagtgg ggtcccatca 180 aggttcagtg gcagtggatc tgggacagat
ttcactctca ccatcagcag tctgcaacct 240 gaagattttg caacttacta
ctgtcaacag agttacagta cccctccgct caccttcggc 300 caagggacac
aactggagat taaa 324 <210> SEQ ID NO 58 <211> LENGTH:
108 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 58 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg
Ala Ser Gln Ser Ile Asn Asn Tyr 20 25 30 Leu Asn Trp Tyr Gln Gln
Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Thr Ala Ser
Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly
Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80
Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85
90 95 Leu Thr Phe Gly Gln Gly Thr Gln Leu Glu Ile Lys 100 105
<210> SEQ ID NO 59 <211> LENGTH: 18 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 59
cagagcatta acaactat 18 <210> SEQ ID NO 60 <211> LENGTH:
6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 60 Gln Ser Ile Asn Asn Tyr 1 5 <210>
SEQ ID NO 61 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 61
actgcatcc 9 <210> SEQ ID NO 62 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 62 Thr Ala Ser 1 <210> SEQ ID NO 63
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 63 caacagagtt
acagtacccc tccgctcacc 30 <210> SEQ ID NO 64 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 64 Gln Gln Ser Tyr Ser Thr Pro Pro
Leu Thr 1 5 10 <210> SEQ ID NO 65 <211> LENGTH: 366
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 65 gaggtgcagc tggtggagtc tgggggaggc
ttggtacagc ctggggggtc cctgagactc 60 tcctgtggag cctctggatt
caccttcagg aactacgaca tgcactgggt ccgccaaatt 120 acaggaaaag
gtctggagtg ggtctcagct attggtagtg ctggtgacac atactatcca 180
gactccgtga agggccgatt caccatctcc agagaaaatg ccaagaactc cttgtatctt
240 caaatgaaca gcctgagagt cggggacacg gctgtgtatt actgtacaag
agatatccat 300 tgtagtagta ccaggtgcta cggtatggac gtctggggcc
aagggaccac ggtcaccgtc 360 tcctca 366 <210> SEQ ID NO 66
<211> LENGTH: 122 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 66 Glu Val Gln Leu Val
Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg
Leu Ser Cys Gly Ala Ser Gly Phe Thr Phe Arg Asn Tyr 20 25 30 Asp
Met His Trp Val Arg Gln Ile Thr Gly Lys Gly Leu Glu Trp Val 35 40
45 Ser Ala Ile Gly Ser Ala Gly Asp Thr Tyr Tyr Pro Asp Ser Val Lys
50 55 60 Gly Arg Phe Thr Ile Ser Arg Glu Asn Ala Lys Asn Ser Leu
Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Val Gly Asp Thr Ala Val
Tyr Tyr Cys Thr 85 90 95 Arg Asp Ile His Cys Ser Ser Thr Arg Cys
Tyr Gly Met Asp Val Trp 100 105 110 Gly Gln Gly Thr Thr Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 67 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 67 ggattcacct tcaggaacta cgac 24 <210>
SEQ ID NO 68 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 68
Gly Phe Thr Phe Arg Asn Tyr Asp 1 5 <210> SEQ ID NO 69
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 69 attggtagtg
ctggtgacac a 21 <210> SEQ ID NO 70 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 70 Ile Gly Ser Ala Gly Asp Thr 1 5
<210> SEQ ID NO 71 <211> LENGTH: 48 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 71
acaagagata tccattgtag tagtaccagg tgctacggta tggacgtc 48 <210>
SEQ ID NO 72 <211> LENGTH: 16 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 72
Thr Arg Asp Ile His Cys Ser Ser Thr Arg Cys Tyr Gly Met Asp Val 1 5
10 15 <210> SEQ ID NO 73 <211> LENGTH: 324 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 73 gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga
cagagtcacc 60 atcacttgcc gggcaagtca gagcattagc aactatttaa
attggtatca gcagaaacca 120 gggaaagccc ctaagctcct gatctatgct
gcatccagtt tgcaaagtgg ggtcccatca 180 aggttcagtg gcagtggatc
tgggacagat ttcactctca ccatcagcag tctgcaacct 240 gaagattttg
caacttacta ctgtcaacag agttacagta cccctccgat caccttcggc 300
caagggacac gactggagat taaa 324 <210> SEQ ID NO 74 <211>
LENGTH: 108 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 74 Asp Ile Gln Met Thr Gln Ser Pro
Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr
Cys Arg Ala Ser Gln Ser Ile Ser Asn Tyr 20 25 30 Leu Asn Trp Tyr
Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Ala
Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60
Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65
70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr
Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys
100 105 <210> SEQ ID NO 75 <211> LENGTH: 18 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 75 cagagcatta gcaactat 18 <210> SEQ ID NO 76
<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 76 Gln Ser Ile Ser Asn
Tyr 1 5
<210> SEQ ID NO 77 <211> LENGTH: 9 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 77
gctgcatcc 9 <210> SEQ ID NO 78 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 78 Ala Ala Ser 1 <210> SEQ ID NO 79
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 79 caacagagtt
acagtacccc tccgatcacc 30 <210> SEQ ID NO 80 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 80 Gln Gln Ser Tyr Ser Thr Pro Pro
Ile Thr 1 5 10 <210> SEQ ID NO 81 <211> LENGTH: 390
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 81 gaggtgcagc tggtggagtc tgggggaggc
ttggtaaagc ctggggggtc ccttcgactc 60 tcctgtgcag cctctggatt
caaattcagt aatgaatgga tgagctgggt ccgccaggct 120 ccagggaagg
ggctggagtg ggttggccgt attaaaagca aaactgatgg tgggacaaca 180
gactacgctg cacccgtgaa aggcagattc accatctcaa gagatgattc aaaaaatacg
240 ctgtatctgc aaatgaacag cctgaaaacc gaggacacag ccgtgtatta
ctgtaccaca 300 gatcaagatt tttggagtgg ttattatacc ggggctgact
actacggtat ggacgtctgg 360 ggccaaggga ccatggtcac cgtctcctca 390
<210> SEQ ID NO 82 <211> LENGTH: 130 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 82
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Lys Phe Ser Asn
Glu 20 25 30 Trp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Gly Arg Ile Lys Ser Lys Thr Asp Gly Gly Thr
Thr Asp Tyr Ala Ala 50 55 60 Pro Val Lys Gly Arg Phe Thr Ile Ser
Arg Asp Asp Ser Lys Asn Thr 65 70 75 80 Leu Tyr Leu Gln Met Asn Ser
Leu Lys Thr Glu Asp Thr Ala Val Tyr 85 90 95 Tyr Cys Thr Thr Asp
Gln Asp Phe Trp Ser Gly Tyr Tyr Thr Gly Ala 100 105 110 Asp Tyr Tyr
Gly Met Asp Val Trp Gly Gln Gly Thr Met Val Thr Val 115 120 125 Ser
Ser 130 <210> SEQ ID NO 83 <211> LENGTH: 24 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 83 ggattcaaat tcagtaatga atgg 24 <210> SEQ ID NO 84
<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 84 Gly Phe Lys Phe Ser
Asn Glu Trp 1 5 <210> SEQ ID NO 85 <211> LENGTH: 30
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 85 attaaaagca aaactgatgg tgggacaaca 30
<210> SEQ ID NO 86 <211> LENGTH: 10 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 86
Ile Lys Ser Lys Thr Asp Gly Gly Thr Thr 1 5 10 <210> SEQ ID
NO 87 <211> LENGTH: 63 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 87 accacagatc
aagatttttg gagtggttat tataccgggg ctgactacta cggtatggac 60 gtc 63
<210> SEQ ID NO 88 <211> LENGTH: 21 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 88
Thr Thr Asp Gln Asp Phe Trp Ser Gly Tyr Tyr Thr Gly Ala Asp Tyr 1 5
10 15 Tyr Gly Met Asp Val 20 <210> SEQ ID NO 89 <211>
LENGTH: 324 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 89 gacatccaga tgacccagtc tccatcctcc
ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgcc gggcaagtca
gagcattagc agctatttaa attggtatca gcagaaacca 120 gggaaagccc
ctaagctcct gatctatgct gcatccagtt tgcaaagtgg ggtcccatca 180
aggttcagtg gcagtggatc tgggacagat ttcactctca ccatcagcag tctgcaacct
240 gaagattttg caacttacta ctgtcaacag agttacagta cccctccgat
caccttcggc 300 caagggacac gactggagat taaa 324 <210> SEQ ID NO
90 <211> LENGTH: 108 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 90 Asp Ile Gln
Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp
Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr 20 25
30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile
35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe
Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser
Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln
Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly Thr Arg
Leu Glu Ile Lys 100 105 <210> SEQ ID NO 91 <211>
LENGTH: 18 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 91
cagagcatta gcagctat 18 <210> SEQ ID NO 92 <211> LENGTH:
6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 92 Gln Ser Ile Ser Ser Tyr 1 5 <210>
SEQ ID NO 93 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 93
gctgcatcc 9 <210> SEQ ID NO 94 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 94 Ala Ala Ser 1 <210> SEQ ID NO 95
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 95 caacagagtt
acagtacccc tccgatcacc 30 <210> SEQ ID NO 96 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 96 Gln Gln Ser Tyr Ser Thr Pro Pro
Ile Thr 1 5 10 <210> SEQ ID NO 97 <211> LENGTH: 348
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 97 cagatgcagc tccaacagtg gggcgcagga
ctattgaagc cttcggagac cctgtccctc 60 acctgcgttg tctatggtgg
gtccctcaat ggatactatt ggagctggat ccgccagtcc 120 cccgggaagg
ggctggagtg gattggggaa atcgatcata gtggaagcac caactacaac 180
ccgtccctca agaatcgagt caccatgtca gtagacacgt ctaagattca gttctccctg
240 aaactgacct ctgtgaccgt cgcggacacg gctgtgtatt actgtgcgag
agaaggatta 300 ttaccctttg actattgggg ccagggaacc ctggtcaccg tctcctca
348 <210> SEQ ID NO 98 <211> LENGTH: 116 <212>
TYPE: PRT <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 98 Gln Met Gln Leu Gln Gln Trp Gly Ala Gly Leu Leu Lys
Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Val Val Tyr Gly Gly
Ser Leu Asn Gly Tyr 20 25 30 Tyr Trp Ser Trp Ile Arg Gln Ser Pro
Gly Lys Gly Leu Glu Trp Ile 35 40 45 Gly Glu Ile Asp His Ser Gly
Ser Thr Asn Tyr Asn Pro Ser Leu Lys 50 55 60 Asn Arg Val Thr Met
Ser Val Asp Thr Ser Lys Ile Gln Phe Ser Leu 65 70 75 80 Lys Leu Thr
Ser Val Thr Val Ala Asp Thr Ala Val Tyr Tyr Cys Ala 85 90 95 Arg
Glu Gly Leu Leu Pro Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105
110 Thr Val Ser Ser 115 <210> SEQ ID NO 99 <211>
LENGTH: 24 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 99 ggtgggtccc tcaatggata ctat 24
<210> SEQ ID NO 100 <211> LENGTH: 8 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 100
Gly Gly Ser Leu Asn Gly Tyr Tyr 1 5 <210> SEQ ID NO 101
<211> LENGTH: 21 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 101 atcgatcata
gtggaagcac c 21 <210> SEQ ID NO 102 <211> LENGTH: 7
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 102 Ile Asp His Ser Gly Ser Thr 1 5
<210> SEQ ID NO 103 <211> LENGTH: 30 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 103
gcgagagaag gattattacc ctttgactat 30 <210> SEQ ID NO 104
<211> LENGTH: 10 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 104 Ala Arg Glu Gly
Leu Leu Pro Phe Asp Tyr 1 5 10 <210> SEQ ID NO 105
<211> LENGTH: 324 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 105 gaaattgtgt
tgacgcagtc tccaggcacc ctgtctttgt ctccagggga aagagtcacc 60
ctctcctgca gggccagtca gagtgtttac agcaactact tagcctggta ccagcagaat
120 cctggccagg ctcccaggct cctcatctat gctgcatcca acagggccac
tggcatccca 180 gacaggttca gtggcagtgg gtctgggaca gacttcactc
tcaccatcag cagactggag 240 cctgaagatt ttgcggtgta ttactgtcat
cagtatgcta cctcaccttg gacgttcggc 300 caagggacca aggtggaaat caaa 324
<210> SEQ ID NO 106 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 106
Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5
10 15 Glu Arg Val Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Tyr Ser
Asn 20 25 30 Tyr Leu Ala Trp Tyr Gln Gln Asn Pro Gly Gln Ala Pro
Arg Leu Leu 35 40 45 Ile Tyr Ala Ala Ser Asn Arg Ala Thr Gly Ile
Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr
Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr
Tyr Cys His Gln Tyr Ala Thr Ser Pro 85 90 95 Trp Thr Phe Gly Gln
Gly Thr Lys Val Glu Ile Lys
100 105 <210> SEQ ID NO 107 <211> LENGTH: 21
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 107 cagagtgttt acagcaacta c 21 <210>
SEQ ID NO 108 <211> LENGTH: 7 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 108
Gln Ser Val Tyr Ser Asn Tyr 1 5 <210> SEQ ID NO 109
<211> LENGTH: 9 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 109 gctgcatcc 9
<210> SEQ ID NO 110 <211> LENGTH: 3 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 110
Ala Ala Ser 1 <210> SEQ ID NO 111 <211> LENGTH: 27
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 111 catcagtatg ctacctcacc ttggacg 27
<210> SEQ ID NO 112 <211> LENGTH: 9 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 112
His Gln Tyr Ala Thr Ser Pro Trp Thr 1 5 <210> SEQ ID NO 113
<211> LENGTH: 369 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 113 cagctgcagc
tgcaggagtc gggcccagat ctggtgaagc cttcggatac cctgtccctc 60
acctgcactg tctctgatga ctccatcagc agtactactt actactgggc ctggatccgc
120 cagcccccag ggaaggggct ggaatggatt ggcagtatgt cttataatgg
gaacaactac 180 tacaacccgt ccctcaagag tcgagtcgcc atatccgcag
gcacgtccca gaaacagttc 240 tccctgaaac tgacctctgt gactgccgca
gacacggctg tttatcactg tgcgagacat 300 cttggatata acggcaactg
gtaccccttt gacttctggg gccagggaat tctggtcacc 360 gtctcctct 369
<210> SEQ ID NO 114 <211> LENGTH: 123 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 114
Gln Leu Gln Leu Gln Glu Ser Gly Pro Asp Leu Val Lys Pro Ser Asp 1 5
10 15 Thr Leu Ser Leu Thr Cys Thr Val Ser Asp Asp Ser Ile Ser Ser
Thr 20 25 30 Thr Tyr Tyr Trp Ala Trp Ile Arg Gln Pro Pro Gly Lys
Gly Leu Glu 35 40 45 Trp Ile Gly Ser Met Ser Tyr Asn Gly Asn Asn
Tyr Tyr Asn Pro Ser 50 55 60 Leu Lys Ser Arg Val Ala Ile Ser Ala
Gly Thr Ser Gln Lys Gln Phe 65 70 75 80 Ser Leu Lys Leu Thr Ser Val
Thr Ala Ala Asp Thr Ala Val Tyr His 85 90 95 Cys Ala Arg His Leu
Gly Tyr Asn Gly Asn Trp Tyr Pro Phe Asp Phe 100 105 110 Trp Gly Gln
Gly Ile Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 115
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 115 gatgactcca
tcagcagtac tacttactac 30 <210> SEQ ID NO 116 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 116 Asp Asp Ser Ile Ser Ser Thr Thr
Tyr Tyr 1 5 10 <210> SEQ ID NO 117 <211> LENGTH: 21
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 117 atgtcttata atgggaacaa c 21 <210>
SEQ ID NO 118 <211> LENGTH: 7 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 118
Met Ser Tyr Asn Gly Asn Asn 1 5 <210> SEQ ID NO 119
<211> LENGTH: 45 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 119 gcgagacatc
ttggatataa cggcaactgg tacccctttg acttc 45 <210> SEQ ID NO 120
<211> LENGTH: 15 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 120 Ala Arg His Leu
Gly Tyr Asn Gly Asn Trp Tyr Pro Phe Asp Phe 1 5 10 15 <210>
SEQ ID NO 121 <211> LENGTH: 324 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 121
gaaattgtgt tgacgcagtc tccaggcacc ctgtctttgt ctccaggaga aagagccacc
60 ctctcctgca gggccagtca gagtgttagt agtagttatt tagcctggta
ccagcagaaa 120 cctggccagg ctcccaggct cctcatctat ggtgcatcca
gcaggaccac tggcatccca 180 gacaggttca gtggcagtgg gtctgggaca
gacttcactc tcaccatcag cagactggag 240 cctgaagatt ttgcagtgta
ttactgtcag cagtatggta gctcaccttg gacgttcggc 300 caagggacca
aggtggaaat caaa 324 <210> SEQ ID NO 122 <211> LENGTH:
108 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 122 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr
Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg
Ala Ser Gln Ser Val Ser Ser Ser 20 25 30 Tyr Leu Ala Trp Tyr Gln
Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45
Ile Tyr Gly Ala Ser Ser Arg Thr Thr Gly Ile Pro Asp Arg Phe Ser 50
55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu
Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Gly
Ser Ser Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr Lys Val Glu Ile
Lys 100 105 <210> SEQ ID NO 123 <211> LENGTH: 21
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 123 cagagtgtta gtagtagtta t 21 <210>
SEQ ID NO 124 <211> LENGTH: 7 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 124
Gln Ser Val Ser Ser Ser Tyr 1 5 <210> SEQ ID NO 125
<211> LENGTH: 9 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 125 ggtgcatcc 9
<210> SEQ ID NO 126 <211> LENGTH: 3 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 126
Gly Ala Ser 1 <210> SEQ ID NO 127 <211> LENGTH: 27
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 127 cagcagtatg gtagctcacc ttggacg 27
<210> SEQ ID NO 128 <211> LENGTH: 9 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 128
Gln Gln Tyr Gly Ser Ser Pro Trp Thr 1 5 <210> SEQ ID NO 129
<211> LENGTH: 366 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 129 gaagtgcagg
tggtagagtc tgggggcggc ttggtcgagc ctggcaggtc cctgagactc 60
tcctgtaaag cctctggatt cacctttgat gattatgcca tgcactgggt ccgacaaact
120 ccagggaagg ccctggagtg ggtctcgggt attaattgga gtggtaataa
cataggctat 180 gcggactctg tgaagggccg attcaccatc tccaaggacg
acgccaagaa ctccctgtat 240 ctgcaaatga acagtctgag acctgaggac
acggccttat attactgtac aaaagatata 300 agtataactg gaaccctcga
tgcttttgat gtctggggcc aagggacaat ggtcaccgtc 360 tcttca 366
<210> SEQ ID NO 130 <211> LENGTH: 122 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 130
Glu Val Gln Val Val Glu Ser Gly Gly Gly Leu Val Glu Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Lys Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Thr Pro Gly Lys Ala Leu
Glu Trp Val 35 40 45 Ser Gly Ile Asn Trp Ser Gly Asn Asn Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Lys Asp
Asp Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Pro Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Thr Lys Asp Ile Ser
Ile Thr Gly Thr Leu Asp Ala Phe Asp Val Trp 100 105 110 Gly Gln Gly
Thr Met Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 131
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 131 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 132 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 132 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 133 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 133
attaattgga gtggtaataa cata 24 <210> SEQ ID NO 134 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 134 Ile Asn Trp Ser Gly Asn Asn Ile
1 5 <210> SEQ ID NO 135 <211> LENGTH: 45 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 135 acaaaagata taagtataac tggaaccctc gatgcttttg atgtc 45
<210> SEQ ID NO 136 <211> LENGTH: 15 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 136
Thr Lys Asp Ile Ser Ile Thr Gly Thr Leu Asp Ala Phe Asp Val 1 5 10
15 <210> SEQ ID NO 137 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 137 gacatccaga tgacccagtc tccaatttcc gtgtctgcat
ctgtaggaga cagagtcacc 60 atcacttgtc gggcgagtca gggtattagc
aactggttag cctggtatca gcagaaacca 120 gggatagccc ctaaactcct
gatctattct gcatccagtt tacaaagtgg ggtcccatca 180 aggttcagag
gcagtggatc tgggacagac ttcactctca ccatcggcag cctgcagcct 240
gaagattttg caacttacta ttgtcaacag gctcacagtt tcccgctcac tttcggcgga
300 gggaccaagg tggagatcaa a 321 <210> SEQ ID NO 138
<211> LENGTH: 107 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 138
Asp Ile Gln Met Thr Gln Ser Pro Ile Ser Val Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Asn
Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Ile Ala Pro Lys
Leu Leu Ile 35 40 45 Tyr Ser Ala Ser Ser Leu Gln Ser Gly Val Pro
Ser Arg Phe Arg Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu
Thr Ile Gly Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Gln Gln Ala His Ser Phe Pro Leu 85 90 95 Thr Phe Gly Gly Gly
Thr Lys Val Glu Ile Lys 100 105 <210> SEQ ID NO 139
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 139 cagggtatta
gcaactgg 18 <210> SEQ ID NO 140 <211> LENGTH: 6
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 140 Gln Gly Ile Ser Asn Trp 1 5 <210>
SEQ ID NO 141 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 141
tctgcatcc 9 <210> SEQ ID NO 142 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 142 Ser Ala Ser 1 <210> SEQ ID NO 143
<211> LENGTH: 27 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 143 caacaggctc
acagtttccc gctcact 27 <210> SEQ ID NO 144 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 144 Gln Gln Ala His Ser Phe Pro Leu Thr 1 5
<210> SEQ ID NO 145 <211> LENGTH: 348 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 145
caggtgcaat tagtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc
60 tcctgtgcag cgtctggatt caccttcagt agctatggca tgcactgggt
ccgccaggct 120 ccaggcaagg ggctggagtg ggtggcaatt atatggtctg
atggagatag tgaatataat 180 ctagactccg taaagggccg attcaccatc
tccagagaca attccaagaa cacgctgtat 240 ctgcaaatga acagtctgag
agtcgaagac tcggctgtat attactgtgc gagagatcga 300 gaccttgagg
atatctgggg ccaagggaca atggtcaccg tctcttca 348 <210> SEQ ID NO
146 <211> LENGTH: 116 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 146 Gln Val Gln
Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25
30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ala Ile Ile Trp Ser Asp Gly Asp Ser Glu Tyr Asn Leu Asp
Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Val Glu Asp
Ser Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asp Arg Asp Leu Glu Asp
Ile Trp Gly Gln Gly Thr Met Val 100 105 110 Thr Val Ser Ser 115
<210> SEQ ID NO 147 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 147
ggattcacct tcagtagcta tggc 24 <210> SEQ ID NO 148 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 148 Gly Phe Thr Phe Ser Ser Tyr Gly
1 5 <210> SEQ ID NO 149 <211> LENGTH: 24 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 149 atatggtctg atggagatag tgaa 24 <210> SEQ ID NO
150 <211> LENGTH: 8 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 150 Ile Trp Ser
Asp Gly Asp Ser Glu 1 5 <210> SEQ ID NO 151 <211>
LENGTH: 27 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 151 gcgagagatc gagaccttga ggatatc
27 <210> SEQ ID NO 152 <211> LENGTH: 9 <212>
TYPE: PRT <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 152 Ala Arg Asp Arg Asp Leu Glu Asp Ile 1 5 <210>
SEQ ID NO 153 <211> LENGTH: 321 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 153
gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtcggaga cagagtcacc
60 atcacttgcc gggcaagtca gggcattaga aatgatttag gctggtatca
gcagaaacca 120 gggaaagccc ctaagcgcct gatctatgct gcatccaatt
tgcaaagtgg ggtcccatca 180 aggttcagcg gcagtggatc tgggacagag
ttcactctca caatcagcag cctgcagcct 240 gaagattttg caacttatta
ctgtctacag cataatagtt atccgctcac tttcggcgga 300 gggaccaagg
tggagatcaa a 321
<210> SEQ ID NO 154 <211> LENGTH: 107 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 154
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Arg Asn
Asp 20 25 30 Leu Gly Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Arg Leu Ile 35 40 45 Tyr Ala Ala Ser Asn Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Leu Gln His Asn Ser Tyr Pro Leu 85 90 95 Thr Phe Gly Gly Gly
Thr Lys Val Glu Ile Lys 100 105 <210> SEQ ID NO 155
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 155 cagggcatta
gaaatgat 18 <210> SEQ ID NO 156 <211> LENGTH: 6
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 156 Gln Gly Ile Arg Asn Asp 1 5 <210>
SEQ ID NO 157 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 157
gctgcatcc 9 <210> SEQ ID NO 158 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 158 Ala Ala Ser 1 <210> SEQ ID NO 159
<211> LENGTH: 27 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 159 ctacagcata
atagttatcc gctcact 27 <210> SEQ ID NO 160 <211> LENGTH:
9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 160 Leu Gln His Asn Ser Tyr Pro Leu Thr 1 5
<210> SEQ ID NO 161 <211> LENGTH: 351 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 161
gaggtgcagt tgttggagtc tgggggagtt ttggtacagc ctggggggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttagt aattttggca tgacgtgggt
ccgccaggct 120 ccagggaagg gactggagtg ggtctcaggt attagtggtg
gcggtcgtga cacatacttc 180 gcagactccg tgaagggccg gttcaccatc
tccagagaca attccaagaa tacgttgtat 240 ctacagatga acagcctgaa
aggcgaggac acggccgtat attactgtgt gaagtgggga 300 aatatttact
ttgactactg gggccaggga accctggtca ccgtctcatc a 351 <210> SEQ
ID NO 162 <211> LENGTH: 117 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 162 Glu Val Gln
Leu Leu Glu Ser Gly Gly Val Leu Val Gln Pro Gly Gly 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Phe 20 25
30 Gly Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ser Gly Ile Ser Gly Gly Gly Arg Asp Thr Tyr Phe Ala Asp
Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Gly Glu Asp
Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Trp Gly Asn Ile Tyr Phe
Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115
<210> SEQ ID NO 163 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 163
ggattcacct ttagtaattt tggc 24 <210> SEQ ID NO 164 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 164 Gly Phe Thr Phe Ser Asn Phe Gly
1 5 <210> SEQ ID NO 165 <211> LENGTH: 24 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 165 attagtggtg gcggtcgtga caca 24 <210> SEQ ID NO
166 <211> LENGTH: 8 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 166 Ile Ser Gly
Gly Gly Arg Asp Thr 1 5 <210> SEQ ID NO 167 <211>
LENGTH: 30 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 167 gtgaagtggg gaaatattta
ctttgactac 30 <210> SEQ ID NO 168 <211> LENGTH: 10
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 168 Val Lys Trp Gly Asn Ile Tyr Phe Asp Tyr 1
5 10 <210> SEQ ID NO 169 <211> LENGTH: 321 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 169 gacatccaga tgacccagtc tccatcctcc ctgtctgcat
ctgtcggaga cagcatcacc 60 atcacttgcc gggcgagtct gtccattaac
acctttttaa attggtatca gcagaaacca 120 gggaaagccc ctaacctcct
gatctatgct gcgtccagtt tacatggtgg ggtcccatca 180
aggttcagtg gcagcggctc tgggacagat ttcactctca ccatcagaac tcttcaacct
240 gaagattttg caacttacta ctgtcaacag agttccaata ccccattcac
tttcggccct 300 gggaccgtag tggatttcag a 321 <210> SEQ ID NO
170 <211> LENGTH: 107 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 170 Asp Ile Gln
Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp
Ser Ile Thr Ile Thr Cys Arg Ala Ser Leu Ser Ile Asn Thr Phe 20 25
30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Asn Leu Leu Ile
35 40 45 Tyr Ala Ala Ser Ser Leu His Gly Gly Val Pro Ser Arg Phe
Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Arg
Thr Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln
Ser Ser Asn Thr Pro Phe 85 90 95 Thr Phe Gly Pro Gly Thr Val Val
Asp Phe Arg 100 105 <210> SEQ ID NO 171 <211> LENGTH:
18 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 171 ctgtccatta acaccttt 18 <210> SEQ ID
NO 172 <211> LENGTH: 6 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 172 Leu Ser Ile
Asn Thr Phe 1 5 <210> SEQ ID NO 173 <211> LENGTH: 9
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 173 gctgcgtcc 9 <210> SEQ ID NO 174
<211> LENGTH: 3 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 174 Ala Ala Ser 1
<210> SEQ ID NO 175 <211> LENGTH: 27 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 175
caacagagtt ccaatacccc attcact 27 <210> SEQ ID NO 176
<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 176 Gln Gln Ser Ser
Asn Thr Pro Phe Thr 1 5 <210> SEQ ID NO 177 <211>
LENGTH: 363 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 177 gaggtgcagc tggtggagtc
tgggggagga gtggtacggc cgggggggtc cctgagactc 60 tcctgtgcag
cctctggatt cacttttgat gactatggca tgagttgggt ccgccaagtt 120
ccagggaagg ggctggagtg ggtctcaggt attagttgga atgatggtaa gacagtttat
180 gcagagtctg tgaagggccg attcatcatc tccagagaca acgccaagaa
ctccctgtat 240 ctggaaatga atagtctgag agccgaggac acggccttat
attactgtgc gagagattgg 300 cagtacttga tagagcggta ctttgactac
tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210> SEQ ID NO
178 <211> LENGTH: 121 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 178 Glu Val Gln
Leu Val Glu Ser Gly Gly Gly Val Val Arg Pro Gly Gly 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25
30 Gly Met Ser Trp Val Arg Gln Val Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ser Gly Ile Ser Trp Asn Asp Gly Lys Thr Val Tyr Ala Glu
Ser Val 50 55 60 Lys Gly Arg Phe Ile Ile Ser Arg Asp Asn Ala Lys
Asn Ser Leu Tyr 65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp
Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Trp Gln Tyr Leu Ile
Glu Arg Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr
Val Ser Ser 115 120 <210> SEQ ID NO 179 <211> LENGTH:
24 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 179 ggattcactt ttgatgacta tggc 24 <210>
SEQ ID NO 180 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 180
Gly Phe Thr Phe Asp Asp Tyr Gly 1 5 <210> SEQ ID NO 181
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 181 attagttgga
atgatggtaa gaca 24 <210> SEQ ID NO 182 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 182 Ile Ser Trp Asn Asp Gly Lys Thr 1 5
<210> SEQ ID NO 183 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 183
gcgagagatt ggcagtactt gatagagcgg tactttgact ac 42 <210> SEQ
ID NO 184 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 184 Ala Arg Asp
Trp Gln Tyr Leu Ile Glu Arg Tyr Phe Asp Tyr 1 5 10 <210> SEQ
ID NO 185 <211> LENGTH: 324 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 185 gaaatagttt tgacacagag tcccggcaca
ctgtcactct ctcccgggga aagagccacc 60 ttgtcatgta gagcaagtca
gtcagtctct agctcttatc tcgcctggta ccagcagaag 120 ccgggacagg
cccctagact gctgatctac ggggcaagtt ccagggccac cggaatcccc 180
gaccggttca gtggaagcgg aagcggaacc gattttactt tgacgatttc tagactggag
240 ccagaggatt tcgccgttta ctattgtcaa cagtacggaa gcagcccgtg
gacgtttggc 300 cagggcacga aggtagaaat caag 324 <210> SEQ ID NO
186 <211> LENGTH: 108 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 186 Glu Ile Val
Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu
Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser 20 25
30 Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu
35 40 45 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg
Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile
Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln
Gln Tyr Gly Ser Ser Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr Lys
Val Glu Ile Lys 100 105 <210> SEQ ID NO 187 <211>
LENGTH: 21 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 187 cagtcagtct ctagctctta t 21
<210> SEQ ID NO 188 <211> LENGTH: 7 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 188
Gln Ser Val Ser Ser Ser Tyr 1 5 <210> SEQ ID NO 189
<211> LENGTH: 9 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 189 ggggcaagt 9
<210> SEQ ID NO 190 <211> LENGTH: 3 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 190
Gly Ala Ser 1 <210> SEQ ID NO 191 <211> LENGTH: 27
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 191 caacagtacg gaagcagccc gtggacg 27
<210> SEQ ID NO 192 <211> LENGTH: 9 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 192
Gln Gln Tyr Gly Ser Ser Pro Trp Thr 1 5 <210> SEQ ID NO 193
<211> LENGTH: 363 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 193 gaggtgcagc
tggtggagtc tgggggaggt gtggtacggc ctggggggtc cctgagactc 60
tcctgtacag cctctggatt cacctttgat gattatggca tgagctgggt ccgccaagct
120 ccagggaagg ggctggagtg gatctctggt attggttgga ctggtggtcg
gtcaagttat 180 gcagactctg tgaggggccg attcaccatc tccagagaca
acgccaagaa ttccctgtat 240 ctgcaaatga acagtctggg agccgaggac
acggccttgt attattgtgc aagagatcgg 300 cagtggctgg tgcagtggta
ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 194 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 194
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Arg Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Gly Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Ile 35 40 45 Ser Gly Ile Gly Trp Thr Gly Gly Arg Ser Ser
Tyr Ala Asp Ser Val 50 55 60 Arg Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Gly
Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Arg Gln
Trp Leu Val Gln Trp Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 195
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 195 ggattcacct
ttgatgatta tggc 24 <210> SEQ ID NO 196 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 196 Gly Phe Thr Phe Asp Asp Tyr Gly 1 5
<210> SEQ ID NO 197 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 197
attggttgga ctggtggtcg gtca 24 <210> SEQ ID NO 198 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 198 Ile Gly Trp Thr Gly Gly Arg Ser
1 5 <210> SEQ ID NO 199 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 199 gcaagagatc ggcagtggct ggtgcagtgg tactttgact ac 42
<210> SEQ ID NO 200 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 200 Ala Arg Asp Arg Gln Trp Leu Val Gln Trp
Tyr Phe Asp Tyr 1 5 10 <210> SEQ ID NO 201 <211>
LENGTH: 324 <212> TYPE: DNA <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 201 gacatccaga tgacccagtc
tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgcc
gggcaagtca gagcattagc agctatttaa attggtatca gcagaaacca 120
gggaaagccc ctaagctcct gatctatgct gcatccagtt tgcaaagtgg ggtcccgtca
180 aggttcagtg gcagtggatc tgggacagat ttcactctca ccatcagcag
tctgcaacct 240 gaagattttg caacttacta ctgtcaacag agttacagta
cccctccgat caccttcggc 300 caagggacac gactggagat taaa 324
<210> SEQ ID NO 202 <211> LENGTH: 108 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 202
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5
10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser
Tyr 20 25 30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro
Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu
Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr
Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln
Gly Thr Arg Leu Glu Ile Lys 100 105 <210> SEQ ID NO 203
<211> LENGTH: 18 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 203 cagagcatta
gcagctat 18 <210> SEQ ID NO 204 <211> LENGTH: 6
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 204 Gln Ser Ile Ser Ser Tyr 1 5 <210>
SEQ ID NO 205 <211> LENGTH: 9 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 205
gctgcatcc 9 <210> SEQ ID NO 206 <211> LENGTH: 3
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 206 Ala Ala Ser 1 <210> SEQ ID NO 207
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 207 caacagagtt
acagtacccc tccgatcacc 30 <210> SEQ ID NO 208 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 208 Gln Gln Ser Tyr Ser Thr Pro Pro
Ile Thr 1 5 10 <210> SEQ ID NO 209 <211> LENGTH: 363
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 209 gaggtgcagc tggtggagtc tgggggaaga
gtggtacggc cgggggggtc cctgagactc 60 tcctgtgcag cctctggatt
cacttttgat gactatggca tgagttgggt ccgccaactt 120 ccagggaagg
gcctggagtg ggtcgcaggt attagttgga atgatggtaa gacagtttat 180
gcagagtctg tgaagggccg attcatcatc tccagagaca acgccaagaa ctccctgcat
240 ctggagatga acagtctgag agccgaggac acggccttat attactgtgc
gcgagattgg 300 caatacttaa tagatcgtta ctttgacttc tggggtcagg
gaaccctggt caccgtctcc 360 tca 363 <210> SEQ ID NO 210
<211> LENGTH: 121 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 210 Glu Val Gln Leu
Val Glu Ser Gly Gly Arg Val Val Arg Pro Gly Gly 1 5 10 15 Ser Leu
Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30
Gly Met Ser Trp Val Arg Gln Leu Pro Gly Lys Gly Leu Glu Trp Val 35
40 45 Ala Gly Ile Ser Trp Asn Asp Gly Lys Thr Val Tyr Ala Glu Ser
Val 50 55 60 Lys Gly Arg Phe Ile Ile Ser Arg Asp Asn Ala Lys Asn
Ser Leu His 65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp Thr
Ala Leu Tyr Tyr Cys 85 90 95 Ala Arg Asp Trp Gln Tyr Leu Ile Asp
Arg Tyr Phe Asp Phe Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 211 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 211 ggattcactt ttgatgacta tggc 24 <210>
SEQ ID NO 212 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 212
Gly Phe Thr Phe Asp Asp Tyr Gly 1 5 <210> SEQ ID NO 213
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 213 attagttgga
atgatggtaa gaca 24 <210> SEQ ID NO 214 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 214 Ile Ser Trp Asn Asp Gly Lys Thr 1 5
<210> SEQ ID NO 215 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE:
215
gcgcgagatt ggcaatactt aatagatcgt tactttgact tc 42 <210> SEQ
ID NO 216 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 216 Ala Arg Asp
Trp Gln Tyr Leu Ile Asp Arg Tyr Phe Asp Phe 1 5 10 <210> SEQ
ID NO 217 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 217 gaagtgcagc
tggtggagtc tgggggaggc ttggtgcagc ctggcgggtc cctgagactc 60
tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct
120 ccagggaagg gcctggagtg ggtctcaggt attggttgga gtagtggtag
cataggctat 180 gcggactctg tgaagggccg attcaccatc tccagagaca
acgccaagaa ctccttgtat 240 ctgcaaatgg acagtctgag acctgaggac
tcagccttat attactgtgc aaaagcctat 300 acatttatga ttaccctcta
ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 218 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 218
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Gly Trp Ser Ser Gly Ser Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asp Ser Leu Arg
Pro Glu Asp Ser Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Ala Tyr Thr
Phe Met Ile Thr Leu Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 219
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 219 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 220 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 220 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 221 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 221
attggttgga gtagtggtag cata 24 <210> SEQ ID NO 222 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 222 Ile Gly Trp Ser Ser Gly Ser Ile
1 5 <210> SEQ ID NO 223 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 223 gcaaaagcct atacatttat gattaccctc tactttgact ac 42
<210> SEQ ID NO 224 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 224
Ala Lys Ala Tyr Thr Phe Met Ile Thr Leu Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 225 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 225
gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgat gattatgaca tgcactgggt
ccggcaagct 120 ccagggaagg gcctggagtg ggtgtcaggg agtggttgga
ataggggtag tttaggctat 180 gcggattctg tgaagggccg attcaccatc
tccagagaca acgccaagaa gtccctgtat 240 ctgcaaatga acagtgtgag
agttgaggac acggccttgt attactgtgc aaaaggcttt 300 gtagtggtat
cagctgctta ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 226 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 226
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Asp Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ser Gly Trp Asn Arg Gly Ser Leu Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Lys Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Val Arg
Val Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Gly Phe Val
Val Val Ser Ala Ala Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 227
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 227 ggattcacct
ttgatgatta tgac 24 <210> SEQ ID NO 228 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 228 Gly Phe Thr Phe Asp Asp Tyr Asp 1 5
<210> SEQ ID NO 229 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 229
agtggttgga ataggggtag ttta 24 <210> SEQ ID NO 230 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic
<400> SEQUENCE: 230 Ser Gly Trp Asn Arg Gly Ser Leu 1 5
<210> SEQ ID NO 231 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 231
gcaaaaggct ttgtagtggt atcagctgct tactttgact ac 42 <210> SEQ
ID NO 232 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 232 Ala Lys Gly
Phe Val Val Val Ser Ala Ala Tyr Phe Asp Tyr 1 5 10 <210> SEQ
ID NO 233 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 233 caggtgcagc
tggtgcagtc tggggctgag gtgaagaggc ctgggtcctc ggtgaaggtc 60
tcctgcaagg tatctggagt caccttcagg aattttgcta tcatctgggt gcgacaggcc
120 cctggacaag ggcttgagtg gatgggagga atcatccctt tctttagtgc
agcaaattac 180 gcacagagct tccagggcag agtcacgatt accccggacg
aatccacgag cacagccttc 240 atggagctgg ccagtctgag atctgaggac
acggccgttt attattgtgc gagagagggg 300 gaacgtggac acacctatgg
gtttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 234 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 234
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Arg Pro Gly Ser 1 5
10 15 Ser Val Lys Val Ser Cys Lys Val Ser Gly Val Thr Phe Arg Asn
Phe 20 25 30 Ala Ile Ile Trp Val Arg Gln Ala Pro Gly Gln Gly Leu
Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Phe Phe Ser Ala Ala Asn
Tyr Ala Gln Ser Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Pro Asp
Glu Ser Thr Ser Thr Ala Phe 65 70 75 80 Met Glu Leu Ala Ser Leu Arg
Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Glu Gly Glu
Arg Gly His Thr Tyr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 235
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 235 ggagtcacct
tcaggaattt tgct 24 <210> SEQ ID NO 236 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 236 Gly Val Thr Phe Arg Asn Phe Ala 1 5
<210> SEQ ID NO 237 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 237
atcatccctt tctttagtgc agca 24 <210> SEQ ID NO 238 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 238 Ile Ile Pro Phe Phe Ser Ala Ala
1 5 <210> SEQ ID NO 239 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 239 gcgagagagg gggaacgtgg acacacctat gggtttgact ac 42
<210> SEQ ID NO 240 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 240
Ala Arg Glu Gly Glu Arg Gly His Thr Tyr Gly Phe Asp Tyr 1 5 10
<210> SEQ ID NO 241 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 241
gaagtgcagc tggtggagtc tgggggaggc ttggtacagt ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt
ccgacaacct 120 ccagggaagg gcctggaatg ggtctcaggt attaactgga
atagaggtag gacaggctat 180 gcggactctg tgaagggccg attcaccatc
tccagagaca acgccaagaa ctccctgtat 240 ctgcaaatga acgatctgag
agttgaggat acggccttgt attactgtgc aaaagccgaa 300 cagtggctgg
acgagggata ctttgactac tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 242 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 242
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Ser Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Pro Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Asn Trp Asn Arg Gly Arg Thr Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Asp Leu Arg
Val Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Ala Glu Gln
Trp Leu Asp Glu Gly Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 243
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 243 ggattcacct
ttgatgatta tgcc 24 <210> SEQ ID NO 244 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 244 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5
<210> SEQ ID NO 245 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 245 attaactgga atagaggtag gaca 24 <210>
SEQ ID NO 246 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 246
Ile Asn Trp Asn Arg Gly Arg Thr 1 5 <210> SEQ ID NO 247
<211> LENGTH: 42 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 247 gcaaaagccg
aacagtggct ggacgaggga tactttgact ac 42 <210> SEQ ID NO 248
<211> LENGTH: 14 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 248 Ala Lys Ala Glu
Gln Trp Leu Asp Glu Gly Tyr Phe Asp Tyr 1 5 10 <210> SEQ ID
NO 249 <211> LENGTH: 363 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 249 gaggtgcagc
tggtggagtc tgggggaggc ttggtgcagc ggggggggtc cctgagactc 60
tcctgtgcag cctctggatt cagctttagc agctatgcca tgaactgggt ccgccaggct
120 ccagggaagg ggctggagtg ggtctcaact attagtgata gtggtggtag
tacatactac 180 gcagactccg tgaagggccg gttcaccatt tccagagaca
attccaagaa cacgctgtct 240 ctgcaaatga acagcctgag agccgaggac
acggccgtat attactgtgc gaaagatcag 300 ggtgggagtt acccctacta
ctttcactac tggggccagg gaaccctggt caccgtctcc 360 tca 363 <210>
SEQ ID NO 250 <211> LENGTH: 121 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 250
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Arg Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Ser Phe Ser Ser
Tyr 20 25 30 Ala Met Asn Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Thr Ile Ser Asp Ser Gly Gly Ser Thr Tyr
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ser Lys Asn Thr Leu Ser 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Gln Gly
Gly Ser Tyr Pro Tyr Tyr Phe His Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 251
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 251 ggattcagct
ttagcagcta tgcc 24 <210> SEQ ID NO 252 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 252 Gly Phe Ser Phe Ser Ser Tyr Ala 1 5
<210> SEQ ID NO 253 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 253
attagtgata gtggtggtag taca 24 <210> SEQ ID NO 254 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 254 Ile Ser Asp Ser Gly Gly Ser Thr
1 5 <210> SEQ ID NO 255 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 255 gcgaaagatc agggtgggag ttacccctac tactttcact ac 42
<210> SEQ ID NO 256 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 256
Ala Lys Asp Gln Gly Gly Ser Tyr Pro Tyr Tyr Phe His Tyr 1 5 10
<210> SEQ ID NO 257 <211> LENGTH: 363 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 257
gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttgag gattatgcca tgcactgggt
ccggcaagct 120 ccagggaagg gcctggagtg ggtctcaggt attggttgga
gtaatgtaaa gataggctat 180 gcggactctg tgaagggccg attcaccatc
tccagagaca atgtcaggaa ctccctatat 240 ctgcaaatga acagtctgag
aactgaggac acggccttct attactgtgt aaaagcctat 300 acatctatgc
ttaccctcta ctttgactat tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 258 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 258
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Glu Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Gly Ile Gly Trp Ser Asn Val Lys Ile Gly
Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Val Arg Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Thr Glu Asp Thr Ala Phe Tyr Tyr Cys 85 90 95 Val Lys Ala Tyr Thr
Ser Met Leu Thr Leu Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 259
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 259 ggattcacct
ttgaggatta tgcc 24 <210> SEQ ID NO 260 <211> LENGTH:
8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 260 Gly Phe Thr Phe Glu Asp Tyr Ala 1 5
<210> SEQ ID NO 261 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 261
attggttgga gtaatgtaaa gata 24 <210> SEQ ID NO 262 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 262 Ile Gly Trp Ser Asn Val Lys Ile
1 5 <210> SEQ ID NO 263 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 263 gtaaaagcct atacatctat gcttaccctc tactttgact at 42
<210> SEQ ID NO 264 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 264
Val Lys Ala Tyr Thr Ser Met Leu Thr Leu Tyr Phe Asp Tyr 1 5 10
<210> SEQ ID NO 265 <211> LENGTH: 354 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 265
caggtgcagc tggtgcagtc tggggctgag gtgaagaggc ctggggcctc agtgaaggtt
60 tcctgcaagg catctggata caccttcacc agcttctata tgtactgggt
gcgacaggcc 120 cctggacaag ggcttgagtg gatgggaata atcaacccta
gtgatggtag cacaagcaac 180 gcacagaagt tccagggcag agtcaccatg
accagggaca cgtccacgag tacagtctac 240 atggagctga gcagcctgag
atctgaggac acggccgtgt attactgtgc gagacgggtg 300 gctggggata
tttttgatat ctggggccaa gggacaatgg tcaccgtctc ttca 354 <210>
SEQ ID NO 266 <211> LENGTH: 118 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 266
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Arg Pro Gly Ala 1 5
10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser
Phe 20 25 30 Tyr Met Tyr Trp Val Arg Gln Ala Pro Gly Gln Gly Leu
Glu Trp Met 35 40 45 Gly Ile Ile Asn Pro Ser Asp Gly Ser Thr Ser
Asn Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Arg Asp
Thr Ser Thr Ser Thr Val Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg
Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Arg Val Ala
Gly Asp Ile Phe Asp Ile Trp Gly Gln Gly Thr 100 105 110 Met Val Thr
Val Ser Ser 115 <210> SEQ ID NO 267 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 267 ggatacacct tcaccagctt ctat 24 <210>
SEQ ID NO 268 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 268
Gly Tyr Thr Phe Thr Ser Phe Tyr 1 5 <210> SEQ ID NO 269
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 269 atcaacccta
gtgatggtag caca 24 <210> SEQ ID NO 270 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 270 Ile Asn Pro Ser Asp Gly Ser Thr 1 5
<210> SEQ ID NO 271 <211> LENGTH: 33 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 271
gcgagacggg tggctgggga tatttttgat atc 33 <210> SEQ ID NO 272
<211> LENGTH: 11 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 272 Ala Arg Arg Val
Ala Gly Asp Ile Phe Asp Ile 1 5 10 <210> SEQ ID NO 273
<211> LENGTH: 357 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 273 caggtgcagc
tgcaggagtc gggcccagga ctggtgaagc cttcggagac cctgtccctc 60
acctgcactg tctctggtgg ctccatcagt agttaccact ggaactggat ccggcagagt
120 ccagggaagg gactggaatg gattggatat atctattata ttgggagcac
cgactataat 180 ccctccctcg agagtcgagt caccatatca gtagacacgt
ccaagaacca gttctccctg 240 aagctgagtt ctgtgaccgc tgcggacacg
gccgtgtatt actgtgcgag agtccccgtg 300 ggagctacag gggcttctga
tgtctggggc caagggacaa tggtcaccgt ctcttca 357 <210> SEQ ID NO
274 <211> LENGTH: 119 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 274 Gln Val Gln
Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr
Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr 20 25
30 His Trp Asn Trp Ile Arg Gln Ser Pro Gly Lys Gly Leu Glu Trp Ile
35 40 45 Gly Tyr Ile Tyr Tyr Ile Gly Ser Thr Asp Tyr Asn Pro Ser
Leu Glu 50 55 60 Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn
Gln Phe Ser Leu 65 70 75 80 Lys Leu Ser Ser Val Thr Ala Ala Asp Thr
Ala Val Tyr Tyr Cys Ala 85 90 95 Arg Val Pro Val Gly Ala Thr Gly
Ala Ser Asp Val Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser
Ser 115 <210> SEQ ID NO 275 <211> LENGTH: 24
<212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 275
ggtggctcca tcagtagtta ccac 24 <210> SEQ ID NO 276 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 276 Gly Gly Ser Ile Ser Ser Tyr His
1 5 <210> SEQ ID NO 277 <211> LENGTH: 21 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 277 atctattata ttgggagcac c 21 <210> SEQ ID NO 278
<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 278 Ile Tyr Tyr Ile
Gly Ser Thr 1 5 <210> SEQ ID NO 279 <211> LENGTH: 39
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 279 gcgagagtcc ccgtgggagc tacaggggct
tctgatgtc 39 <210> SEQ ID NO 280 <211> LENGTH: 13
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 280 Ala Arg Val Pro Val Gly Ala Thr Gly Ala
Ser Asp Val 1 5 10 <210> SEQ ID NO 281 <211> LENGTH:
363 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 281 gaggtgcagc tggtggagtc tgggggaagt
gtggttcgac ctggggggtc cctgagactc 60 tcctgtgtag tctctggatt
cacctttgag gattatggtt tgagctgggt ccgccaaatt 120 ccagggaaag
gactggagtg ggtctctggt attagttgga ctggtggtaa cacaggttat 180
gcagactctg tgaagggccg cttcaccatc tccagagaca acgccaagaa ctccctgtat
240 ctgcaaatga acagtctgag agccgaagac acggccctgt atcactgtac
gagagatcga 300 cagtggctga tgcagtggta ttttgactat tggggccagg
gaaccctggt caccgtctcc 360 tca 363 <210> SEQ ID NO 282
<211> LENGTH: 121 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 282 Glu Val Gln Leu
Val Glu Ser Gly Gly Ser Val Val Arg Pro Gly Gly 1 5 10 15 Ser Leu
Arg Leu Ser Cys Val Val Ser Gly Phe Thr Phe Glu Asp Tyr 20 25 30
Gly Leu Ser Trp Val Arg Gln Ile Pro Gly Lys Gly Leu Glu Trp Val 35
40 45 Ser Gly Ile Ser Trp Thr Gly Gly Asn Thr Gly Tyr Ala Asp Ser
Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn
Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr
Ala Leu Tyr His Cys 85 90 95 Thr Arg Asp Arg Gln Trp Leu Met Gln
Trp Tyr Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val
Ser Ser 115 120 <210> SEQ ID NO 283 <211> LENGTH: 24
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 283 ggattcacct ttgaggatta tggt 24 <210>
SEQ ID NO 284 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 284
Gly Phe Thr Phe Glu Asp Tyr Gly 1 5 <210> SEQ ID NO 285
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 285 attagttgga
ctggtggtaa caca 24 <210> SEQ ID NO 286 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 286 Ile Ser Trp Thr Gly Gly Asn Thr 1 5
<210> SEQ ID NO 287 <211> LENGTH: 42 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 287
acgagagatc gacagtggct gatgcagtgg tattttgact at 42 <210> SEQ
ID NO 288 <211> LENGTH: 14 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 288 Thr Arg Asp
Arg Gln Trp Leu Met Gln Trp Tyr Phe Asp Tyr 1 5 10 <210> SEQ
ID NO 289 <211> LENGTH: 357 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 289 caggtgcagc
tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60
tcctgttcag cctctggatt caccttcagt gcctatgcca tgcactgggt ccgccaggct
120 ccaggcaagg ggctggaatg ggtggcagct atctcatatg gtggaagtga
taaatactat 180 gcagactccg tgaagggccg attcaccatc tccagagaca
attccaagaa cacgctatat 240 ctgcaaatga acagcctgag aactgacgac
acggctgtgt attactgtgc gaaatccgct 300 cactggaact tcttctttga
ctactggggc cagggaaccc tggtcactgt ctcctca 357 <210> SEQ ID NO
290 <211> LENGTH: 119 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 290 Gln Val Gln
Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser
Leu Arg Leu Ser Cys Ser Ala Ser Gly Phe Thr Phe Ser Ala Tyr 20 25
30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val
35 40 45 Ala Ala Ile Ser Tyr Gly Gly Ser Asp Lys Tyr Tyr Ala Asp
Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Thr Leu Tyr 65 70 75 80
Leu Gln Met Asn Ser Leu Arg Thr Asp Asp Thr Ala Val Tyr Tyr Cys 85
90 95 Ala Lys Ser Ala His Trp Asn Phe Phe Phe Asp Tyr Trp Gly Gln
Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> SEQ ID
NO 291 <211> LENGTH: 24 <212> TYPE: DNA <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 291 ggattcacct
tcagtgccta tgcc 24 <210> SEQ ID NO 292 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 292 Gly Phe Thr Phe Ser Ala Tyr Ala 1 5
<210> SEQ ID NO 293 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 293
atctcatatg gtggaagtga taaa 24 <210> SEQ ID NO 294 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 294 Ile Ser Tyr Gly Gly Ser Asp Lys
1 5 <210> SEQ ID NO 295 <211> LENGTH: 36 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 295 gcgaaatccg ctcactggaa cttcttcttt gactac 36
<210> SEQ ID NO 296 <211> LENGTH: 12 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 296
Ala Lys Ser Ala His Trp Asn Phe Phe Phe Asp Tyr 1 5 10 <210>
SEQ ID NO 297 <211> LENGTH: 363 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 297
gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc
60 tcctgtgtag cctctggatt cgcccttcat gattatgcca tgcactgggt
ccggcaagtt 120 ccagggaagg gcctggagtg ggtctcaagt attagttgga
atagtggtgt cataggctat 180 gcggactctc tgaagggccg cttcaccatc
tccagagaca acgccaagaa ctccctgtat 240 ctgcaaatga acagtctgag
agcagaggac acggccttat actactgtgc aaaaggtagt 300 gggagctact
acgtcagttg gttcgacccc tggggccagg gaaccctggt caccgtctcc 360 tca 363
<210> SEQ ID NO 298 <211> LENGTH: 121 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 298
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5
10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Phe Ala Leu His Asp
Tyr 20 25 30 Ala Met His Trp Val Arg Gln Val Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45 Ser Ser Ile Ser Trp Asn Ser Gly Val Ile Gly
Tyr Ala Asp Ser Leu 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Gly Ser Gly
Ser Tyr Tyr Val Ser Trp Phe Asp Pro Trp Gly 100 105 110 Gln Gly Thr
Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 299
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 299 ggattcgccc
ttcatgatta tgcc 24 <210> SEQ ID NO 300 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 300 Gly Phe Ala Leu His Asp Tyr Ala 1 5
<210> SEQ ID NO 301 <211> LENGTH: 24 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 301
attagttgga atagtggtgt cata 24 <210> SEQ ID NO 302 <211>
LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 302 Ile Ser Trp Asn Ser Gly Val Ile
1 5 <210> SEQ ID NO 303 <211> LENGTH: 42 <212>
TYPE: DNA <213> ORGANISM: Artificial Sequence <220>
FEATURE: <223> OTHER INFORMATION: Synthetic <400>
SEQUENCE: 303 gcaaaaggta gtgggagcta ctacgtcagt tggttcgacc cc 42
<210> SEQ ID NO 304 <211> LENGTH: 14 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 304
Ala Lys Gly Ser Gly Ser Tyr Tyr Val Ser Trp Phe Asp Pro 1 5 10
<210> SEQ ID NO 305 <211> LENGTH: 369 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 305
cagctgcagc tgcaggagtc gggcccagga ctggttcagc cttcggagac cctgtccctc
60 acctgcactg tctctggtga ctccatcagt agtactgctt accactggga
ctggatccgc 120 cagccccccg ggaagggact ggagtggatt gggaccatca
cttataatgg gaacacctac 180 ttcaacccgt ccctcaagag tcgagtcacc
atatccgttg acacgtccaa gaaccagttc 240 tccctgaagc tactctctat
gaccgccgca gaaacggctg ttttttactg tgcgcgacat 300 ctaggatata
acagtgactt ctttcccttt gacttctggg gccagggaac cctggtcact 360
gtctcctca 369 <210> SEQ ID NO 306 <211> LENGTH: 123
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 306
Gln Leu Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Gln Pro Ser Glu 1 5
10 15 Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Asp Ser Ile Ser Ser
Thr 20 25 30 Ala Tyr His Trp Asp Trp Ile Arg Gln Pro Pro Gly Lys
Gly Leu Glu 35 40 45 Trp Ile Gly Thr Ile Thr Tyr Asn Gly Asn Thr
Tyr Phe Asn Pro Ser 50 55 60 Leu Lys Ser Arg Val Thr Ile Ser Val
Asp Thr Ser Lys Asn Gln Phe 65 70 75 80 Ser Leu Lys Leu Leu Ser Met
Thr Ala Ala Glu Thr Ala Val Phe Tyr 85 90 95 Cys Ala Arg His Leu
Gly Tyr Asn Ser Asp Phe Phe Pro Phe Asp Phe 100 105 110 Trp Gly Gln
Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> SEQ ID NO 307
<211> LENGTH: 30 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 307 ggtgactcca
tcagtagtac tgcttaccac 30 <210> SEQ ID NO 308 <211>
LENGTH: 10 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
Synthetic <400> SEQUENCE: 308 Gly Asp Ser Ile Ser Ser Thr Ala
Tyr His 1 5 10 <210> SEQ ID NO 309 <211> LENGTH: 21
<212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 309 atcacttata atgggaacac c 21 <210>
SEQ ID NO 310 <211> LENGTH: 7 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 310
Ile Thr Tyr Asn Gly Asn Thr 1 5 <210> SEQ ID NO 311
<211> LENGTH: 45 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 311 gcgcgacatc
taggatataa cagtgacttc tttccctttg acttc 45 <210> SEQ ID NO 312
<211> LENGTH: 15 <212> TYPE: PRT <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 312 Ala Arg His Leu
Gly Tyr Asn Ser Asp Phe Phe Pro Phe Asp Phe 1 5 10 15 <210>
SEQ ID NO 313 <211> LENGTH: 357 <212> TYPE: DNA
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 313
gaggtgcagc tggtggagtc tgggggaggc ctggtacggc cgggggggtc cctgagactc
60 tcctgtgcag cctctggatt cacctttagc acctatgcca tggcctgggt
ccgccagact 120 ccagggaagg ggctggaggg ggtctcagct attgggggta
gtggtgatag tacctattat 180 gtcgactccg tgaagggccg gttcaccatc
tccagggaca actccaagag cacgcttttt 240 ctgcaaatga atagcctgag
agccgaggac acggccgttt attactgtgt gaaagtccgg 300 aattacgacg
gttcttttga tatctggggc caagggacaa tggtcaccgt ctcttca 357 <210>
SEQ ID NO 314 <211> LENGTH: 119 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 314
Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Arg Pro Gly Gly 1 5
10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Thr
Tyr 20 25 30 Ala Met Ala Trp Val Arg Gln Thr Pro Gly Lys Gly Leu
Glu Gly Val 35 40 45 Ser Ala Ile Gly Gly Ser Gly Asp Ser Thr Tyr
Tyr Val Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp
Asn Ser Lys Ser Thr Leu Phe 65 70 75 80 Leu Gln Met Asn Ser Leu Arg
Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Val Arg Asn
Tyr Asp Gly Ser Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val
Thr Val Ser Ser 115 <210> SEQ ID NO 315 <211> LENGTH:
24 <212> TYPE: DNA <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 315 ggattcacct ttagcaccta tgcc 24 <210>
SEQ ID NO 316 <211> LENGTH: 8 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 316
Gly Phe Thr Phe Ser Thr Tyr Ala 1 5 <210> SEQ ID NO 317
<211> LENGTH: 24 <212> TYPE: DNA <213> ORGANISM:
Artificial Sequence <220> FEATURE: <223> OTHER
INFORMATION: Synthetic <400> SEQUENCE: 317 attgggggta
gtggtgatag tacc 24 <210> SEQ ID NO 318 <211> LENGTH: 8
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: Synthetic
<400> SEQUENCE: 318 Ile Gly Gly Ser Gly Asp Ser Thr 1 5
<210> SEQ ID NO 319 <211> LENGTH: 36 <212> TYPE:
DNA <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: Synthetic <400> SEQUENCE: 319
gtgaaagtcc ggaattacga cggttctttt gatatc 36 <210> SEQ ID NO
320 <211> LENGTH: 12 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: Synthetic <400> SEQUENCE: 320 Val Lys Val
Arg Asn Tyr Asp Gly Ser Phe Asp Ile 1 5 10 <210> SEQ ID NO
321 <211> LENGTH: 174 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE: <223>
OTHER INFORMATION: hPD-1-MMH <220> FEATURE: <223> OTHER
INFORMATION: 1-146: aa 25-170 of NP_005009.2 with C93S <220>
FEATURE: <223> OTHER INFORMATION: 147-174:
myc-myc-hexahistidine <400> SEQUENCE: 321 Leu Asp Ser Pro Asp
Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Val
Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser
Asn Thr Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro
35 40 45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg
Ser Gln 50 55 60 Pro Gly Gln Asp Ser Arg Phe Arg Val Thr Gln Leu
Pro Asn Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg
Arg Asn Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu
Ala Pro Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu
Arg Val Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro
Ser Pro Ser Pro Arg Pro Ala Gly Gln Phe Gln Thr 130 135 140 Leu Val
Glu Gln Lys Leu Ile Ser Glu Glu Asp Leu Gly Gly Glu Gln 145 150 155
160 Lys Leu Ile Ser Glu Glu Asp Leu His His His His His His 165 170
<210> SEQ ID NO 322 <211> LENGTH: 174 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: MfPD-1-MMH <220> FEATURE:
<223> OTHER INFORMATION: 1-146: M.fascicularis PD-1 with C93S
<220> FEATURE: <223> OTHER INFORMATION: 147-174:
myc-myc-hexahistidine <400> SEQUENCE: 322 Leu Glu Ser Pro Asp
Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Leu
Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser
Asn Ala Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40
45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln
50 55 60 Pro Gly Arg Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn
Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn
Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro
Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val
Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro
Ser Pro Arg Pro Ala Gly Gln Phe Gln Ala 130 135 140 Leu Val Glu Gln
Lys Leu Ile Ser Glu Glu Asp Leu Gly Gly Glu Gln 145 150 155 160 Lys
Leu Ile Ser Glu Glu Asp Leu His His His His His His 165 170
<210> SEQ ID NO 323 <211> LENGTH: 379 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-1-mFc <220> FEATURE:
<223> OTHER INFORMATION: 1-146: aa 25-170 of NP-005009.2 with
C93S <220> FEATURE: <223> OTHER INFORMATION: 147-379:
mFc: aa 98-330 of P01863 <400> SEQUENCE: 323 Leu Asp Ser Pro
Asp Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu
Val Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30
Ser Asn Thr Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35
40 45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser
Gln 50 55 60 Pro Gly Gln Asp Ser Arg Phe Arg Val Thr Gln Leu Pro
Asn Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg
Asn Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala
Pro Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg
Val Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser
Pro Ser Pro Arg Pro Ala Gly Gln Phe Gln Thr 130 135 140 Leu Val Glu
Pro Arg Gly Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys 145 150 155 160
Cys Pro Ala Pro Asn Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro 165
170 175 Pro Lys Ile Lys Asp Val Leu Met Ile Ser Leu Ser Pro Ile Val
Thr 180 185 190 Cys Val Val Val Asp Val Ser Glu Asp Asp Pro Asp Val
Gln Ile Ser 195 200 205 Trp Phe Val Asn Asn Val Glu Val His Thr Ala
Gln Thr Gln Thr His 210 215 220 Arg Glu Asp Tyr Asn Ser Thr Leu Arg
Val Val Ser Ala Leu Pro Ile 225 230 235 240 Gln His Gln Asp Trp Met
Ser Gly Lys Glu Phe Lys Cys Lys Val Asn 245 250 255 Asn Lys Asp Leu
Pro Ala Pro Ile Glu Arg Thr Ile Ser Lys Pro Lys 260 265 270 Gly Ser
Val Arg Ala Pro Gln Val Tyr Val Leu Pro Pro Pro Glu Glu 275 280 285
Glu Met Thr Lys Lys Gln Val Thr Leu Thr Cys Met Val Thr Asp Phe 290
295 300 Met Pro Glu Asp Ile Tyr Val Glu Trp Thr Asn Asn Gly Lys Thr
Glu 305 310 315 320 Leu Asn Tyr Lys Asn Thr Glu Pro Val Leu Asp Ser
Asp Gly Ser Tyr 325 330 335 Phe Met Tyr Ser Lys Leu Arg Val Glu Lys
Lys Asn Trp Val Glu Arg 340 345 350 Asn Ser Tyr Ser Cys Ser Val Val
His Glu Gly Leu His Asn His His 355 360 365 Thr Thr Lys Ser Phe Ser
Arg Thr Pro Gly Lys 370 375 <210> SEQ ID NO 324 <211>
LENGTH: 373 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
hPD-1-hFc <220> FEATURE: <223> OTHER INFORMATION:
1-146: aa 25-170 of NP_005009.2 with C93S <220> FEATURE:
<223> OTHER INFORMATION: 147-373: hFc: aa 104-330 of P01857
<400> SEQUENCE: 324 Leu Asp Ser Pro Asp Arg Pro Trp Asn Pro
Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Val Val Thr Glu Gly Asp
Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser Asn Thr Ser Glu Ser
Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40 45 Ser Asn Gln Thr
Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln 50 55 60 Pro Gly
Gln Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn Gly Arg 65 70 75 80
Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn Asp Ser Gly Thr 85
90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro Lys Ala Gln Ile Lys
Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val Thr Glu Arg Arg Ala
Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro Ser Pro Arg Pro Ala
Gly Gln Phe Gln Thr 130 135 140 Leu Val Asp Lys Thr His Thr Cys Pro
Pro Cys Pro Ala Pro Glu Leu 145 150 155 160 Leu Gly Gly Pro Ser Val
Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr 165 170 175 Leu Met Ile Ser
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val 180 185 190 Ser His
Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val 195 200 205
Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser 210
215 220 Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp
Leu 225 230 235 240 Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys
Ala Leu Pro Ala 245 250 255 Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys
Gly Gln Pro Arg Glu Pro 260 265 270 Gln Val Tyr Thr Leu Pro Pro Ser
Arg Asp Glu Leu Thr Lys Asn Gln 275 280 285 Val Ser Leu Thr Cys Leu
Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala 290 295 300 Val Glu Trp Glu
Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr 305 310 315 320 Pro
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu 325 330
335 Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser
340 345 350 Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser
Leu Ser 355 360 365 Leu Ser Pro Gly Lys 370 <210> SEQ ID NO
325 <211> LENGTH: 448 <212> TYPE: PRT <213>
ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-L1-hFc <220> FEATURE:
<223> OTHER INFORMATION: 1-221: aa 19-239 of NP_054862.1
<220> FEATURE: <223> OTHER INFORMATION: 222-448: hFc:
aa 104-330 of P01857 <400> SEQUENCE: 325 Phe Thr Val Thr Val
Pro Lys Asp Leu Tyr Val Val Glu Tyr Gly Ser 1 5 10 15 Asn Met Thr
Ile Glu Cys Lys Phe Pro Val Glu Lys Gln Leu Asp Leu 20 25 30 Ala
Ala Leu Ile Val Tyr Trp Glu Met Glu Asp Lys Asn Ile Ile Gln 35 40
45 Phe Val His Gly Glu Glu Asp Leu Lys Val Gln His Ser Ser Tyr Arg
50 55 60 Gln Arg Ala Arg Leu Leu Lys Asp Gln Leu Ser Leu Gly Asn
Ala Ala 65 70 75 80 Leu Gln Ile Thr Asp Val Lys Leu Gln Asp Ala Gly
Val Tyr Arg Cys 85 90 95 Met Ile Ser Tyr Gly Gly Ala Asp Tyr Lys
Arg Ile Thr Val Lys Val 100 105 110 Asn Ala Pro Tyr Asn Lys Ile Asn
Gln Arg Ile Leu Val Val Asp Pro 115 120 125 Val Thr Ser Glu His Glu
Leu Thr Cys Gln Ala Glu Gly Tyr Pro Lys 130 135 140 Ala Glu Val Ile
Trp Thr Ser Ser Asp His Gln Val Leu Ser Gly Lys 145 150 155 160 Thr
Thr Thr Thr Asn Ser Lys Arg Glu Glu Lys Leu Phe Asn Val Thr 165 170
175 Ser Thr Leu Arg Ile Asn Thr Thr Thr Asn Glu Ile Phe Tyr Cys Thr
180 185 190 Phe Arg Arg Leu Asp Pro Glu Glu Asn His Thr Ala Glu Leu
Val Ile 195 200 205 Pro Glu Leu Pro Leu Ala His Pro Pro Asn Glu Arg
Thr Asp Lys Thr 210 215 220 His Thr Cys Pro Pro Cys Pro Ala Pro Glu
Leu Leu Gly Gly Pro Ser 225 230 235 240 Val Phe Leu Phe Pro Pro Lys
Pro Lys Asp Thr Leu Met Ile Ser Arg 245 250 255 Thr Pro Glu Val Thr
Cys Val Val Val Asp Val Ser His Glu Asp Pro 260 265 270 Glu Val Lys
Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala 275 280 285 Lys
Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val 290 295
300 Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr
305 310 315 320 Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile
Glu Lys Thr 325 330 335 Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu 340 345 350 Pro Pro Ser Arg Asp Glu Leu Thr Lys
Asn Gln Val Ser Leu Thr Cys 355 360 365 Leu Val Lys Gly Phe Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ser 370 375 380 Asn Gly Gln Pro Glu
Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp 385 390 395 400 Ser Asp
Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser 405 410 415
Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala 420
425 430 Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys 435 440 445 <210> SEQ ID NO 326 <211> LENGTH: 454
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: hPD-L1-mFc
<220> FEATURE: <223> OTHER INFORMATION: 1-221: aa
19-239 of NP_054862.1 <220> FEATURE: <223> OTHER
INFORMATION: 222-454: mFc: aa 98-330 of P01863 <400>
SEQUENCE: 326 Phe Thr Val Thr Val Pro Lys Asp Leu Tyr Val Val Glu
Tyr Gly Ser 1 5 10 15 Asn Met Thr Ile Glu Cys Lys Phe Pro Val Glu
Lys Gln Leu Asp Leu 20 25 30 Ala Ala Leu Ile Val Tyr Trp Glu Met
Glu Asp Lys Asn Ile Ile Gln 35 40 45 Phe Val His Gly Glu Glu Asp
Leu Lys Val Gln His Ser Ser Tyr Arg 50 55 60 Gln Arg Ala Arg Leu
Leu Lys Asp Gln Leu Ser Leu Gly Asn Ala Ala 65 70 75 80 Leu Gln Ile
Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr Arg Cys 85 90 95 Met
Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile Thr Val Lys Val 100 105
110 Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg Ile Leu Val Val Asp Pro
115 120 125 Val Thr Ser Glu His Glu Leu Thr Cys Gln Ala Glu Gly Tyr
Pro Lys 130 135 140 Ala Glu Val Ile Trp Thr Ser Ser Asp His Gln Val
Leu Ser Gly Lys 145 150 155 160 Thr Thr Thr Thr Asn Ser Lys Arg Glu
Glu Lys Leu Phe Asn Val Thr 165 170 175 Ser Thr Leu Arg Ile Asn Thr
Thr Thr Asn Glu Ile Phe Tyr Cys Thr 180 185 190 Phe Arg Arg Leu Asp
Pro Glu Glu Asn His Thr Ala Glu Leu Val Ile 195 200 205 Pro Glu Leu
Pro Leu Ala His Pro Pro Asn Glu Arg Thr Glu Pro Arg 210 215 220 Gly
Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys Cys Pro Ala Pro Asn 225 230
235 240 Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Ile Lys
Asp 245 250 255 Val Leu Met Ile Ser Leu Ser Pro Ile Val Thr Cys Val
Val Val Asp 260 265 270 Val Ser Glu Asp Asp Pro Asp Val Gln Ile Ser
Trp Phe Val Asn Asn 275 280 285 Val Glu Val His Thr Ala Gln Thr Gln
Thr His Arg Glu Asp Tyr Asn 290 295 300 Ser Thr Leu Arg Val Val Ser
Ala Leu Pro Ile Gln His Gln Asp Trp 305 310 315 320 Met Ser Gly Lys
Glu Phe Lys Cys Lys Val Asn Asn Lys Asp Leu Pro 325 330 335 Ala Pro
Ile Glu Arg Thr Ile Ser Lys Pro Lys Gly Ser Val Arg Ala 340 345 350
Pro Gln Val Tyr Val Leu Pro Pro Pro Glu Glu Glu Met Thr Lys Lys 355
360 365 Gln Val Thr Leu Thr Cys Met Val Thr Asp Phe Met Pro Glu Asp
Ile 370 375 380 Tyr Val Glu Trp Thr Asn Asn Gly Lys Thr Glu Leu Asn
Tyr Lys Asn 385 390 395 400 Thr Glu Pro Val Leu Asp Ser Asp Gly Ser
Tyr Phe Met Tyr Ser Lys 405 410 415 Leu Arg Val Glu Lys Lys Asn Trp
Val Glu Arg Asn Ser Tyr Ser Cys 420 425 430 Ser Val Val His Glu Gly
Leu His Asn His His Thr Thr Lys Ser Phe 435 440 445 Ser Arg Thr Pro
Gly Lys 450 <210> SEQ ID NO 327 <211> LENGTH: 288
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: hPD-1
NP_005009.2 <400> SEQUENCE: 327 Met Gln Ile Pro Gln Ala Pro
Trp Pro Val Val Trp Ala Val Leu Gln 1 5 10 15 Leu Gly Trp Arg Pro
Gly Trp Phe Leu Asp Ser Pro Asp Arg Pro Trp 20 25 30 Asn Pro Pro
Thr Phe Ser Pro Ala Leu Leu Val Val Thr Glu Gly Asp 35 40 45 Asn
Ala Thr Phe Thr Cys Ser Phe Ser Asn Thr Ser Glu Ser Phe Val 50 55
60 Leu Asn Trp Tyr Arg Met Ser Pro Ser Asn Gln Thr Asp Lys Leu Ala
65 70 75 80 Ala Phe Pro Glu Asp Arg Ser Gln Pro Gly Gln Asp Cys Arg
Phe Arg 85 90 95 Val Thr Gln Leu Pro Asn Gly Arg Asp Phe His Met
Ser Val Val Arg 100 105 110 Ala Arg Arg Asn Asp Ser Gly Thr Tyr Leu
Cys Gly Ala Ile Ser Leu 115 120 125 Ala Pro Lys Ala Gln Ile Lys Glu
Ser Leu Arg Ala Glu Leu Arg Val 130 135 140 Thr Glu Arg Arg Ala Glu
Val Pro Thr Ala His Pro Ser Pro Ser Pro 145 150 155 160 Arg Pro Ala
Gly Gln Phe Gln Thr Leu Val Val Gly Val Val Gly Gly 165 170 175 Leu
Leu Gly Ser Leu Val Leu Leu Val Trp Val Leu Ala Val Ile Cys 180 185
190 Ser Arg Ala Ala Arg Gly Thr Ile Gly Ala Arg Arg Thr Gly Gln Pro
195 200 205 Leu Lys Glu Asp Pro Ser Ala Val Pro Val Phe Ser Val Asp
Tyr Gly 210 215 220 Glu Leu Asp Phe Gln Trp Arg Glu Lys Thr Pro Glu
Pro Pro Val Pro 225 230 235 240 Cys Val Pro Glu Gln Thr Glu Tyr Ala
Thr Ile Val Phe Pro Ser Gly 245 250 255 Met Gly Thr Ser Ser Pro Ala
Arg Arg Gly Ser Ala Asp Gly Pro Arg 260 265 270 Ser Ala Gln Pro Leu
Arg Pro Glu Asp Gly His Cys Ser Trp Pro Leu 275 280 285 <210>
SEQ ID NO 328 <211> LENGTH: 290 <212> TYPE: PRT
<213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: hPD-L1 NP_054862.1 <400>
SEQUENCE: 328
Met Arg Ile Phe Ala Val Phe Ile Phe Met Thr Tyr Trp His Leu Leu 1 5
10 15 Asn Ala Phe Thr Val Thr Val Pro Lys Asp Leu Tyr Val Val Glu
Tyr 20 25 30 Gly Ser Asn Met Thr Ile Glu Cys Lys Phe Pro Val Glu
Lys Gln Leu 35 40 45 Asp Leu Ala Ala Leu Ile Val Tyr Trp Glu Met
Glu Asp Lys Asn Ile 50 55 60 Ile Gln Phe Val His Gly Glu Glu Asp
Leu Lys Val Gln His Ser Ser 65 70 75 80 Tyr Arg Gln Arg Ala Arg Leu
Leu Lys Asp Gln Leu Ser Leu Gly Asn 85 90 95 Ala Ala Leu Gln Ile
Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr 100 105 110 Arg Cys Met
Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile Thr Val 115 120 125 Lys
Val Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg Ile Leu Val Val 130 135
140 Asp Pro Val Thr Ser Glu His Glu Leu Thr Cys Gln Ala Glu Gly Tyr
145 150 155 160 Pro Lys Ala Glu Val Ile Trp Thr Ser Ser Asp His Gln
Val Leu Ser 165 170 175 Gly Lys Thr Thr Thr Thr Asn Ser Lys Arg Glu
Glu Lys Leu Phe Asn 180 185 190 Val Thr Ser Thr Leu Arg Ile Asn Thr
Thr Thr Asn Glu Ile Phe Tyr 195 200 205 Cys Thr Phe Arg Arg Leu Asp
Pro Glu Glu Asn His Thr Ala Glu Leu 210 215 220 Val Ile Pro Glu Leu
Pro Leu Ala His Pro Pro Asn Glu Arg Thr His 225 230 235 240 Leu Val
Ile Leu Gly Ala Ile Leu Leu Cys Leu Gly Val Ala Leu Thr 245 250 255
Phe Ile Phe Arg Leu Arg Lys Gly Arg Met Met Asp Val Lys Lys Cys 260
265 270 Gly Ile Gln Asp Thr Asn Ser Lys Lys Gln Ser Asp Thr His Leu
Glu 275 280 285 Glu Thr 290 <210> SEQ ID NO 329 <211>
LENGTH: 379 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
aa1-146: M.fascicularis PD-1 (with C93S change) aa147-379: mFc tag
(aa 98-330 of P01863) <400> SEQUENCE: 329 Leu Glu Ser Pro Asp
Arg Pro Trp Asn Pro Pro Thr Phe Ser Pro Ala 1 5 10 15 Leu Leu Leu
Val Thr Glu Gly Asp Asn Ala Thr Phe Thr Cys Ser Phe 20 25 30 Ser
Asn Ala Ser Glu Ser Phe Val Leu Asn Trp Tyr Arg Met Ser Pro 35 40
45 Ser Asn Gln Thr Asp Lys Leu Ala Ala Phe Pro Glu Asp Arg Ser Gln
50 55 60 Pro Gly Arg Asp Ser Arg Phe Arg Val Thr Gln Leu Pro Asn
Gly Arg 65 70 75 80 Asp Phe His Met Ser Val Val Arg Ala Arg Arg Asn
Asp Ser Gly Thr 85 90 95 Tyr Leu Cys Gly Ala Ile Ser Leu Ala Pro
Lys Ala Gln Ile Lys Glu 100 105 110 Ser Leu Arg Ala Glu Leu Arg Val
Thr Glu Arg Arg Ala Glu Val Pro 115 120 125 Thr Ala His Pro Ser Pro
Ser Pro Arg Pro Ala Gly Gln Phe Gln Ala 130 135 140 Leu Val Glu Pro
Arg Gly Pro Thr Ile Lys Pro Cys Pro Pro Cys Lys 145 150 155 160 Cys
Pro Ala Pro Asn Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro 165 170
175 Pro Lys Ile Lys Asp Val Leu Met Ile Ser Leu Ser Pro Ile Val Thr
180 185 190 Cys Val Val Val Asp Val Ser Glu Asp Asp Pro Asp Val Gln
Ile Ser 195 200 205 Trp Phe Val Asn Asn Val Glu Val His Thr Ala Gln
Thr Gln Thr His 210 215 220 Arg Glu Asp Tyr Asn Ser Thr Leu Arg Val
Val Ser Ala Leu Pro Ile 225 230 235 240 Gln His Gln Asp Trp Met Ser
Gly Lys Glu Phe Lys Cys Lys Val Asn 245 250 255 Asn Lys Asp Leu Pro
Ala Pro Ile Glu Arg Thr Ile Ser Lys Pro Lys 260 265 270 Gly Ser Val
Arg Ala Pro Gln Val Tyr Val Leu Pro Pro Pro Glu Glu 275 280 285 Glu
Met Thr Lys Lys Gln Val Thr Leu Thr Cys Met Val Thr Asp Phe 290 295
300 Met Pro Glu Asp Ile Tyr Val Glu Trp Thr Asn Asn Gly Lys Thr Glu
305 310 315 320 Leu Asn Tyr Lys Asn Thr Glu Pro Val Leu Asp Ser Asp
Gly Ser Tyr 325 330 335 Phe Met Tyr Ser Lys Leu Arg Val Glu Lys Lys
Asn Trp Val Glu Arg 340 345 350 Asn Ser Tyr Ser Cys Ser Val Val His
Glu Gly Leu His Asn His His 355 360 365 Thr Thr Lys Ser Phe Ser Arg
Thr Pro Gly Lys 370 375 <210> SEQ ID NO 330 <211>
LENGTH: 444 <212> TYPE: PRT <213> ORGANISM: Artificial
Sequence <220> FEATURE: <223> OTHER INFORMATION:
H4H7798N aa1-117: HCVR aa118-444: HC constant <400> SEQUENCE:
330 Glu Val Gln Leu Leu Glu Ser Gly Gly Val Leu Val Gln Pro Gly Gly
1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser
Asn Phe 20 25 30 Gly Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly
Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Gly Gly Gly Arg Asp Thr
Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg
Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu
Lys Gly Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Trp Gly
Asn Ile Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr
Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125
Ala Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys 130
135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn
Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala
Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val
Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Lys Thr Tyr Thr Cys
Asn Val Asp His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg
Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro 210 215 220 Pro Cys Pro Ala
Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe 225 230 235 240 Pro
Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val 245 250
255 Thr Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe
260 265 270 Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr
Lys Pro 275 280 285 Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val
Ser Val Leu Thr 290 295 300 Val Leu His Gln Asp Trp Leu Asn Gly Lys
Glu Tyr Lys Cys Lys Val 305 310 315 320 Ser Asn Lys Gly Leu Pro Ser
Ser Ile Glu Lys Thr Ile Ser Lys Ala 325 330 335 Lys Gly Gln Pro Arg
Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln 340 345 350 Glu Glu Met
Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly 355 360 365 Phe
Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro 370 375
380 Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser
385 390 395 400 Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg
Trp Gln Glu 405 410 415 Gly Asn Val Phe Ser Cys Ser Val Met His Glu
Ala Leu His Asn His 420 425 430 Tyr Thr Gln Lys Ser Leu Ser Leu Ser
Leu Gly Lys 435 440 <210> SEQ ID NO 331 <211> LENGTH:
214 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: H4H7798N
aa1-107: LCVR aa108-214: LC constant <400> SEQUENCE: 331 Asp
Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10
15 Asp Ser Ile Thr Ile Thr Cys Arg Ala Ser Leu Ser Ile Asn Thr Phe
20 25 30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Asn Leu
Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu His Gly Gly Val Pro Ser
Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr
Ile Arg Thr Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys
Gln Gln Ser Ser Asn Thr Pro Phe 85 90 95 Thr Phe Gly Pro Gly Thr
Val Val Asp Phe Arg Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe
Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala
Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135
140
Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145
150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser
Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys
His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser
Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210
<210> SEQ ID NO 332 <211> LENGTH: 449 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H7795N2 aa1-122: HCVR aa123-449:
HC constant <400> SEQUENCE: 332 Glu Val Gln Val Val Glu Ser
Gly Gly Gly Leu Val Glu Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser
Cys Lys Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His
Trp Val Arg Gln Thr Pro Gly Lys Ala Leu Glu Trp Val 35 40 45 Ser
Gly Ile Ser Trp Ser Gly Asn Asn Ile Gly Tyr Ala Asp Ser Val 50 55
60 Lys Gly Arg Phe Thr Ile Ser Lys Asp Asp Ala Lys Asn Ser Leu Tyr
65 70 75 80 Leu Gln Met Asn Ser Leu Arg Pro Glu Asp Thr Ala Leu Tyr
Tyr Cys 85 90 95 Thr Lys Asp Ile Ser Ile Thr Gly Thr Leu Asp Ala
Phe Asp Val Trp 100 105 110 Gly Gln Gly Thr Met Val Thr Val Ser Ser
Ala Ser Thr Lys Gly Pro 115 120 125 Ser Val Phe Pro Leu Ala Pro Cys
Ser Arg Ser Thr Ser Glu Ser Thr 130 135 140 Ala Ala Leu Gly Cys Leu
Val Lys Asp Tyr Phe Pro Glu Pro Val Thr 145 150 155 160 Val Ser Trp
Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 175 Ala
Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185
190 Val Pro Ser Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp
195 200 205 His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser
Lys Tyr 210 215 220 Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe
Leu Gly Gly Pro 225 230 235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro
Lys Asp Thr Leu Met Ile Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys
Val Val Val Asp Val Ser Gln Glu Asp 260 265 270 Pro Glu Val Gln Phe
Asn Trp Tyr Val Asp Gly Val Glu Val His Asn 275 280 285 Ala Lys Thr
Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val 290 295 300 Val
Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu 305 310
315 320 Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu
Lys 325 330 335 Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
Val Tyr Thr 340 345 350 Leu Pro Pro Ser Gln Glu Glu Met Thr Lys Asn
Gln Val Ser Leu Thr 355 360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser
Asp Ile Ala Val Glu Trp Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn
Asn Tyr Lys Thr Thr Pro Pro Val Leu 385 390 395 400 Asp Ser Asp Gly
Ser Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys 405 410 415 Ser Arg
Trp Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu 420 425 430
Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly 435
440 445 Lys <210> SEQ ID NO 333 <211> LENGTH: 214
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: H4H7795N2
aa1-107: LCVR aa108-214: LC constant <400> SEQUENCE: 333 Asp
Ile Gln Met Thr Gln Ser Pro Ile Ser Val Ser Ala Ser Val Gly 1 5 10
15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Asn Trp
20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Ile Ala Pro Lys Leu
Leu Ile 35 40 45 Tyr Ser Ala Ser Ser Leu Gln Ser Gly Val Pro Ser
Arg Phe Arg Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr
Ile Gly Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys
Gln Gln Ala His Ser Phe Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe
Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala
Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140
Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145
150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser
Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys
His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser
Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210
<210> SEQ ID NO 334 <211> LENGTH: 446 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H9008P aa1-119: HCVR aa120-446: HC
constant <400> SEQUENCE: 334 Glu Val Gln Leu Val Glu Ser Gly
Gly Gly Leu Val Arg Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys
Ala Ala Ser Gly Phe Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ala Trp
Val Arg Gln Thr Pro Gly Lys Gly Leu Glu Gly Val 35 40 45 Ser Ala
Ile Gly Gly Ser Gly Asp Ser Thr Tyr Tyr Val Asp Ser Val 50 55 60
Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Ser Thr Leu Phe 65
70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr
Tyr Cys 85 90 95 Val Lys Val Arg Asn Tyr Asp Gly Ser Phe Asp Ile
Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser Ala Ser Thr
Lys Gly Pro Ser Val Phe 115 120 125 Pro Leu Ala Pro Cys Ser Arg Ser
Thr Ser Glu Ser Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp
Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly
Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln
Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185
190 Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro
195 200 205 Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly
Pro Pro 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu Gly Gly
Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr
Leu Met Ile Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val
Asp Val Ser Gln Glu Asp Pro Glu Val 260 265 270 Gln Phe Asn Trp Tyr
Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg
Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu
Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310
315 320 Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile
Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr
Leu Pro Pro 340 345 350 Ser Gln Glu Glu Met Thr Lys Asn Gln Val Ser
Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala
Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys
Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe
Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Glu
Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430
Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys 435 440 445
<210> SEQ ID NO 335 <211> LENGTH: 215 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H9008P aa1-108: LCVR aa109-215: LC
constant <400> SEQUENCE: 335 Glu Ile Val Leu Thr Gln Ser Pro
Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15
Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser 20
25 30 Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu
Leu 35 40 45 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp
Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr
Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys
Gln Gln Tyr Gly Ser Ser Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr
Lys Val Glu Ile Lys Arg Thr Val Ala 100 105 110 Ala Pro Ser Val Phe
Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser 115 120 125 Gly Thr Ala
Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu 130 135 140 Ala
Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser 145 150
155 160 Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser
Leu 165 170 175 Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys
His Lys Val 180 185 190 Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser
Ser Pro Val Thr Lys 195 200 205 Ser Phe Asn Arg Gly Glu Cys 210 215
<210> SEQ ID NO 336 <211> LENGTH: 448 <212> TYPE:
PRT <213> ORGANISM: Artificial Sequence <220> FEATURE:
<223> OTHER INFORMATION: H4H9048P2 aa1-121: HCVR aa122-448:
HC constant <400> SEQUENCE: 336 Gln Val Gln Leu Val Gln Ser
Gly Ala Glu Val Lys Arg Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser
Cys Lys Val Ser Gly Val Thr Phe Arg Asn Phe 20 25 30 Ala Ile Ile
Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly
Gly Ile Ile Pro Phe Phe Ser Ala Ala Asn Tyr Ala Gln Ser Phe 50 55
60 Gln Gly Arg Val Thr Ile Thr Pro Asp Glu Ser Thr Ser Thr Ala Phe
65 70 75 80 Met Glu Leu Ala Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr
Tyr Cys 85 90 95 Ala Arg Glu Gly Glu Arg Gly His Thr Tyr Gly Phe
Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser Ala
Ser Thr Lys Gly Pro Ser 115 120 125 Val Phe Pro Leu Ala Pro Cys Ser
Arg Ser Thr Ser Glu Ser Thr Ala 130 135 140 Ala Leu Gly Cys Leu Val
Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 145 150 155 160 Ser Trp Asn
Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala 165 170 175 Val
Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 180 185
190 Pro Ser Ser Ser Leu Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His
195 200 205 Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Ser Lys
Tyr Gly 210 215 220 Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe Leu
Gly Gly Pro Ser 225 230 235 240 Val Phe Leu Phe Pro Pro Lys Pro Lys
Asp Thr Leu Met Ile Ser Arg 245 250 255 Thr Pro Glu Val Thr Cys Val
Val Val Asp Val Ser Gln Glu Asp Pro 260 265 270 Glu Val Gln Phe Asn
Trp Tyr Val Asp Gly Val Glu Val His Asn Ala 275 280 285 Lys Thr Lys
Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val 290 295 300 Ser
Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr 305 310
315 320 Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys
Thr 325 330 335 Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val
Tyr Thr Leu 340 345 350 Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln
Val Ser Leu Thr Cys 355 360 365 Leu Val Lys Gly Phe Tyr Pro Ser Asp
Ile Ala Val Glu Trp Glu Ser 370 375 380 Asn Gly Gln Pro Glu Asn Asn
Tyr Lys Thr Thr Pro Pro Val Leu Asp 385 390 395 400 Ser Asp Gly Ser
Phe Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser 405 410 415 Arg Trp
Gln Glu Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala 420 425 430
Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys 435
440 445 <210> SEQ ID NO 337 <211> LENGTH: 215
<212> TYPE: PRT <213> ORGANISM: Artificial Sequence
<220> FEATURE: <223> OTHER INFORMATION: H4H9048P2
aa1-108: LCVR aa109-215: LC constant <400> SEQUENCE: 337 Asp
Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10
15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr
20 25 30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu
Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser
Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr
Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys
Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Ile Thr Phe Gly Gln Gly
Thr Arg Leu Glu Ile Lys Arg Thr Val Ala 100 105 110 Ala Pro Ser Val
Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser 115 120 125 Gly Thr
Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu 130 135 140
Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser 145
150 155 160 Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr
Ser Leu 165 170 175 Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu
Lys His Lys Val 180 185 190 Tyr Ala Cys Glu Val Thr His Gln Gly Leu
Ser Ser Pro Val Thr Lys 195 200 205 Ser Phe Asn Arg Gly Glu Cys 210
215
* * * * *